2nd Edition Introductory Biology Version 3
2nd Edition Introductory Biology
Student Manual
Copyright Information
Information
Copyright
Introductory Biology Lab Manual
© 2013 eScience Labs, LLC. All rights reserved. This material may not be reproduced, displayed, modified, or distributed, in whole or in part, without the express prior written permission of eScience Labs. Appropriate citation(s) must accompany all excerpts and/or quotations.
For written permissions, please contact info@eScienceLabs.com
Note, educational institutions and customers who have purchased a complete lab kit may reproduce the manual as a print copy for academic use provided that all copies include the following statement: “© 2013 eScience Labs, LLC. All rights reserved.”.
This manual was typeset in 11 Arial and 12 Chalet-London 1960. Arial font provided by Microsoft Office Suite, 2010. Chalet-London 1960 font licensed from House Industries, 2011.
The experiments included within this lab manual are suitable for supervised or unsupervised learning environments. eScience Labs assumes full liability for the safety and techniques employed within this manual provided that all users adhere to the safety guidelines outlined in the mandatory eScience Labs Safety Video, Preface, and Appendix. All users must understand and agree to the eScience Labs safety guidelines prior to beginning their lab experiments. eScience Labs does not condone use of the lab materials provided in its lab kits for any use outside of the curriculum expressly outlined within the lab manual.
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Table of Contents
Introduction
Lab 1
Introduction to Science
Lab 2
General Lab Safety
Lab 3
Chemical Bonding Fundamentals
Lab 4
Introduction to the Microscope
Biological Processes
Lab 5
The Chemistry of Life
Lab 6
Diffusion
Lab 7
Osmosis
Lab 8
Enzymes
Lab 9
Cellular Respiration
Cellular Fundamentals
Lab 10 Cell Structure and Function
Lab 11 Mitosis
Lab 12 Meiosis
Lab 13 DNA and RNA
Lab 14 Mendelian Genetics
Lab 15 Population Genetics
Kingdoms of Life
Lab 16 Taxonomy
Lab 17 Bacteria and Archaea
Lab 18 Protista
Lab 19 Fungi
5
Table of Contents
Plant Kingdom
Lab 20 Energy and Photosynthesis
Lab 21 Plant Circulation
Lab 22 Plant Reproduction
Animal Kingdom
Lab 23 Invertebrates and Vertebrates
Lab 24 Animal Structure
Lab 25 The Circulatory and Respiratory Systems
Lab 26 The Sensory and Nervous Systems
Environmental Biology
Lab 27 Ecology of Organisms
Lab 28 Ecological Interactions
Appendix: Good Lab Techniques
6
Time and Materials
If you are allergic to nitrile, please contact eScience Labs at info@esciencelabs.com and we will send you an alternative type of safety gloves. Swim-caps and rubber bands include latex. Always wear your nitrile safety gloves when working with these items!
Please note that the times listed are approximations and may differ. Please read through the procedure and plan accordingly.
Lab 1 Introduction to Science
Time Required: 1 hour
Additional Materials: None
Lab 2 General Lab Safety
Time Required: 1 hour
Additional Materials: Water
Lab 3 Chemical Bonding Fundamentals
Time Required: 1 hour
Additional Materials: Water, Newspaper, Notebook Paper, Scissors
Lab 4 Introduction to the Microscope
Time Required: 1 hour
Additional Materials: Computer, Internet Access
Lab 5 The Chemistry of Life
Time Required: 2 hours
Additional Materials: Egg White, Hot Pad, Kitchen Knife, Microwave, Paper Towels, Pot (Small), Scissors, Stovetop, Water (Hot and Tap),
7
Time and Materials
Lab 6 Diffusion
Time Required: 2 hours
Additional Materials: Scissors, Stopwatch, Tape, Water
Lab 7 Osmosis
Time Required: 2 1/2 hours (plus 48 hours for preparation and observation)
Additional Materials: Cutting Board, 4 Eggs, Kitchen Knife, 2 Potatoes,
Stopwatch, Water
Lab 8 Enzymes
Time Required: 1 hour (plus 1 hour for preparation)
Additional Materials: Cutting Board, 2 Food Products, Hot Water Bath,
Kitchen Knife, Paper Towel, Saliva Sample, Stopwatch, Water Lab 9 Cellular Respiration
Time Required: 1 hour (plus 24 hours for preparation and time for overnight observations)
Additional Materials: Hot Water, Paper Towels, Stopwatch Lab 10 Cell Structure and Function
Time Required: 1 1/2 hours (plus 24 hours for observation)
Additional Materials: Bowl, 2 Household Items, Warm Water, Computer,
Internet Access Lab 11 Mitosis
Time Required: 3 hours
Additional Materials: None
8
Time and Materials
Lab 12 Meiosis
Time Required: 1 1/2 hours
Additional Materials: Computer, Internet Access Lab 13 DNA and RNA
Time Required: 2 hours
Additional Materials: Fruit, Scissors, Pen/Pencil Lab 14 Mendelian Genetics
Time Required: 1 1/2 hours
Additional Materials: None Lab 15 Population Genetics
Time Required: 1 1/2 hours
Additional Materials: Access to a Printer Lab 16 Taxonomy
Time Required: 1 hour
Additional Materials: None Lab 17 Bacteria and Archaea
Time Required: 1 hour (plus 24 hours preparation and 7 days for observation)
Additional Materials: Microwave or Hot Water Bath, Hot Pad, Refrigerator
9
Time and Materials
Lab 18 Protista
Time Required: 2 hours
Additional Materials: eScience Lab’s Student Portal Account, Internet
Access
Lab 19 Fungi
Time Required: 1 hour (plus 3 - 7 days for observation)
Additional Materials: White Bread, Water Lab 20 Energy and Photosynthesis
Time Required: 1 hour (plus 3 hours for observation)
Additional Materials: Cutting Board, Kitchen Knife, Light Source, Pencil,
Quarter, Scissors, Spinach Leaves (Fresh), Tape, Water Lab 21 Plant Circulation
Time Required: 1 1/2 hours (plus 24 hours for observation)
Additional Materials: 2 Celery Stalks, Cutting Board, Kitchen Knife, Water, Scissors, 3 Additional Environments Lab 22 Plant Reproduction
Time Required: 2 hours
Additional Materials: Fresh Flower (Lily is recommended) Lab 23 Invertebrates and Vertebrates
Time Required: 3 hours
Additional Materials: Paper Towel, Water
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Time and Materials Lab 24 Animal Structure
Time Required: 1 hour (plus 7 additional days for observation)
Additional Materials: 1 Egg, Water
Lab 25 The Circulatory and Respiratory Systems
Time Required: 2 hours
Additional Materials: Water, Scissors
Lab 26 The Sensory and Nervous Systems
Time Required: 2 1/2 hours
Additional Materials: Pen/Pencil, Tape, White Construction or Printer
Paper, Water, Willing Participant
Lab 27 Ecology of Organisms
Time Required: 30 minutes (plus 7 days for observation)
Additional Materials: Paper Towel, Scissors, Sunny Location, Water
Lab 28 Ecological Interactions
Time Required: 30 minutes
Additional Materials: Large Bowl, Paper Towel, Large Pitcher, Water
11
Safety Information
Lab Safety eScience Labs, LLC designs every kit with safety as our top priority. Nonetheless, these are science kits and contain items which must be handled with care.
Safety in the laboratory always comes first!
Always follow the instructions in your laboratory manual and these general rules:
Lab Preparation
•
Please thoroughly read the lab exercise before starting!
•
If you have any doubt as to what you are supposed to be doing and how to do it safely, please STOP and then:
Double-check the manual instructions.
Check www.esciencelabs.com for updates and tips.
Contact us for technical support by phone at 1-888-ESL-Kits (1-888-375-5487) or by email at Help@esciencelabs.com.
•
Read and understand all labels on chemicals.
If you have any questions or concerns, refer to the Material Safely Data Sheets
(MSDS) available at www.esciencelabs.com. The MSDS lists the dangers, storage requirements, exposure treatment and disposal instructions for each chemical.
•
Consult your physician if you are pregnant, allergic to chemicals, or have other medical conditions that may require additional protective measures.
Proper Lab Attire
•
Remove all loose clothing (jackets, sweatshirts, etc.) and always wear closed-toe shoes. •
Long hair should be pulled back and secured and all jewelry (rings, watches, necklaces, earrings, bracelets, etc.) should be removed.
•
Safety glasses or goggles should be worn at all times. In addition, wearing soft contact lenses while conducting experiments is discouraged, as they can absorb potentially harmful chemicals.
•
When handling chemicals, always wear the protective goggles, gloves, and apron provided. 13
Safety Information
Performing the Experiment
•
Do not eat, drink, chew gum, apply cosmetics or smoke while conducting an experiment.
•
Work in a well ventilated area and monitor experiments at all times, unless instructed otherwise. •
When working with chemicals:
Never return unused chemicals to their original container to avoid contamination.
Never place chemicals in an unmarked container to avoid identification or proper disposal problems.
Always put lids back onto chemicals immediately after use to avoid contamination or potential hydration problems.
Never ingest chemicals. If this occurs, seek immediate help.
Call 911 or “Poison Control” 1-800-222-1222
•
Never pipette anything by mouth.
•
Never leave a heat source unattended.
If there is a fire, evacuate the room immediately and dial 911.
Lab Clean-up and Disposal
•
If a spill occurs, consult the MSDS to determine how to clean it up.
•
Never pick up broken glassware with your hands. Use a broom and a dustpan and discard in a safe area.
•
Do not use any part of the lab kit as a container for food.
•
Safely dispose of chemicals. If there are any special requirements for disposal, it will be noted in the lab manual.
•
When finished, wash hands and lab equipment thoroughly with soap and water.
Above all, use common sense! Read the manual carefully. Pay close attention to the safety concerns prior to starting an experiment.
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Student Portal
Introduction:
Concept Animation
ESL Safety Video
ESL Scientific Processes Video
How Big Is It?
Introduction to the Microscope
Measuring Volume Using a
Graduated Cylinder
Unit Conversions
Lab Drill
Biological Processes:
Concept Animation
ESL Biological Processes Video
The Structure of an Atom
Acid/Base Reactions
Diffusion and Osmosis Tutorial
Docking Tutorial
Lab Drill
The Cell:
Concept Animation
Log on to the Student Portal using these easy steps:
Visit our website, www.eScienceLabs.com, and click on the button which says
“Register or Login” on the top right side of the homepage. From here, you will be taken to a login page. If you are registering your kit code for the first time, click the “Create an Account” hyperlink. Locate the kit-code, located on a label on the inside of the kit box lid. Enter this, along with other requested information into the online form to create your user account. Be sure to keep track of your username and password as this is how you will enter the Student Portal for future visits. This establishes your account with the eScience Labs’ Student Portal.
Have fun!
ESL Cell Video
Cell Structure Crossword Puzzle
Interactive Videos of Meiosis
Interactive Videos of Mitosis
Nature’s Review of RNA
DNA Transcription & Translation
How Mutations Work
Riken Center’s Developmental Biology Stem Cell Videos
A Typical Animal Cell
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Student Portal
Construction of the Cell Membrane
The Cell Cycle
Cell Division
DNA Extraction Virtual Lab
Lab Drill
Kingdoms of Life:
Concept Animation
ESL Kingdoms Video
Biology of Bacteria
Biology of Flagellates
Biology of Algae
Protists Under Normal Conditions
Protists With Slowing Agent
Tree of Life Web Project
Virtual Pond Dip
Lab Drill
Plant Kingdom:
Concept Animation
ESL Plant Kingdom Video
Biology of Plants
Lab Drill
Animal Kingdom:
Concept Animation
ESL Animal Kingdom Video
Biology Crossword Puzzle
Biology of Echinoderms
Biology of Cnidarians
Biology of Chordates
Biology of Annelids
Anatomical Terminology: Body Regions
Anatomical Terminology: Relative Position
Regional Body Parts
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Student Portal
The Skelton: Bones & Joints
Major Muscles of the Human Body
The Sense of Hearing
The Sense of Sight
The Sense of Smell
The Sense of Taste
The Anatomy of the Heart
Respiratory Basics
The Respiratory System
Lab Drill
Additional Resources:
Stop Watch
Conversion Tables
17
Sample Labware
19
Lab 1
Introduction to Science
Introduction to Science
Learning Objectives
•
Apply the scientific method, including making observations, developing hypotheses, identifying variables and controls, collecting and analyzing data, and drawing conclusions
•
Use calculations and measurements to connect percent error, significant figures, conversions, accuracy and precision to scientific reasoning
•
Review how to write and format a lab report
Introduction
What is science? You have likely taken several classes throughout your career as a student, and know that it is more than just chapters in a book. Science is a process. It uses evidence to understand the history of the natural world and how it works. Scientific knowledge is not static, but constantly evolving as we understand more about the natural world. Furthermore, the constant development of equipment and techniques allows us to gain an increasingly deeper insight into the natural world. Science begins with observations that can be measured in some way, and often concludes with observations from analyzed data.
Following the scientific method helps to minimize bias and increase validity when testing a theory. It helps scientists collect and organize information in a useful way so that patterns and data can be meaningfully analyzed. As a scientist, you should use the scientific method as you conduct the experiments throughout this manual. Figure 1: The scientific method process.
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Introduction to Science
The process of the scientific method begins with an observation. For example, suppose you observe a plant growing towards a window. This observation could be the first step in designing an experiment! Remember that observations are used to begin the scientific method, but they may also be used to help analyze data.
Observations can be quantitative (measurable), or qualitative (immeasurable; observational). Quantitative observations allow us to record findings as data, and leave little room for subjective error. Qualitative observations cannot be measured. Instead, they rely on human sensory perceptions. The nature of these observations makes them more subjective and susceptible to human error. However, qualitative observations are still able to provide useful information.
For example, suppose you have a handful of pennies. You can make quantitative observations that there are
15 pennies, and each is 1.9 cm in diameter. Both the quantity, and the diameter, can be precisely measured.
You can also make qualitative observations that they are brown, shiny, and smooth. The color and texture are not numerically measured, and may vary based on the individual’s perception or background.
Quantitative observations are generally preferred in science because they involve "hard" data. Because of this, many scientific instruments, such as microscopes and scales, are used to alleviate the need for qualitative observations. Rather than observing that an object is large, scientists can identify specific mass, shapes, structures, etc.
Developing a Hypothesis
Once an observation has been made, the next step is to develop a hypothesis. A hypothesis is a testable statement describing what the scientist thinks will happen in the experiment. In other words, it is a proposed explanation for an event based on observation(s). For every hypothesis, a scientist also develops a null hypothesis. A null hypothesis is a testable statement that, if proven true, means the hypothesis was incorrect.
Both a hypothesis and a null hypothesis must be testable, but only one can be true.
Hypotheses are typically written in an if/then format. For example:
Hypothesis:
If plants are grown in soil with added nutrients, then they will grow faster than plants grown without added nutrients.
Null hypothesis:
If plants are grown in soil with added nutrients, then they will grow at the same rate as plants grown in soil without nutrients.
Figure 2: What affects plant growth?
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Introduction to Science
If plants grow quicker when nutrients are added, then the hypothesis is accepted and the null hypothesis is rejected.
Testing a Hypothesis
There are often many ways to test a hypothesis. However, three rules must always be followed during an experiment for results to be valid.
1. The experiment must be replicable.
2. Only one variable may be tested at a time
3. A control must always be included.
Replicability
Let’s begin by exploring the first rule. Experiments must be replicable to create valid theories. In other words, procedures must always be diligently recorded, and experiments must provide precise. Precise results are those which have very similar values (e.g., 85, 86, and 86.5) over multiple trials. By contrast, accurate results are those which demonstrate what you expected to happen (e.g., you expect amylase, an enzyme found in human saliva, to break down starch molecules during digestion).
Figure 3: Left: Accurate results all hit the bulls-eye on a target. Right: Precise results may not hit the bulls-eye, but they all hit the same region.
The following example demonstrates the significance of experimental replicability. Suppose you conduct an experiment and conclude that ice melts in 30 seconds when placed on a burner, but you do not record your procedure or define the variables. The conclusion that you draw will not be recognized in the scientific community because other scientists cannot repeat your experiment and find the same results. What if another scientist tries to repeat your ice experiment, but does not turn on the burner; or, uses a larger ice chunk than you used? The results will not be the same, because the experiment was not repeated using the same exact procedure. In order for results to be valid, repeated experiments must follow the original experiment exactly. Using this technique, multiple trials performed in this manner should yield comparable results.
25
Introduction to Science
Variables
Variables are defined, measurable components of an experiment. Controlling variables in an experiment allows a scientist to quantify changes that occur. This allows for focused results to be measured; and, for refined conclusions to be drawn. There are two types of variables: independent and dependent variables.
Independent variables are variables that scientists select to change within the experiment. For example, the time of day, amount of substrate, etc. can all be independent variables. Independent variables are also used by scientists to develop hypotheses. The “if” part of the hypothesis describes the independent variable and how the scientist will manipulate it. For example, the independent variable in the hypothesis, “If plants are grown in soil with added nutrients, then they will grow faster than plants grown without added nutrients,” is soil with added nutrients. Experiments can only have one independent variable. In this way, scientists can determine if altering the independent variable is the reason for obtaining a different result. Scientists would not be able to conclusively determine which change affected the data if more than one independent variable is changed in an experiment. However, a hypothesis is usually developed to focus on one variable only. The dependent variable is reflected in the “then” part of the hypothesis. For example, if there is a change in the independent variable, then the dependent variable will also change. Independent variables are always placed on the x-axis of a chart or graph.
Dependent variables are variables which are observed in relationship to the independent variable. Any changes observed in the dependent variable are caused by the changes in the independent variable. In other words, they depend on the independent variable. Common examples of this are: reaction rate, color change, etc. There can be more than one dependent variable in an experiment. Dependent variables are placed on the y-axis of a chart or graph.
Controls
A control is a sample of data collected in an experiment that is not exposed to the independent variable. The control sample reflects the factors that could influence the results of the experiment, but do not reflect the planned changes that might result from manipulating the independent variable. Controls must be identified to eliminate compounding changes that could influence results. Often, the hardest part of designing an experiment is determining how to isolate the independent variable and control all other possible variables. Scientists must be careful not to eliminate or create a factor that could skew the results. For this reason, taking notes to account for unidentified variables is important. This might include factors such as temperature, humidity, time of day, or other environmental conditions that may impact results.
There are two types of controls, positive and negative. Negative controls are data samples in which you expect no change to occur. They help scientists determine that the experimental results are due to the independent variable, rather than an unidentified or unaccounted variable. For example, suppose you need to culture (grow) bacteria and want to include a negative control. You could create this by streaking a sterile loop across an agar plate. Sterile loops should not create any microbial growth; therefore, you expect no change to occur on the agar plate. If no growth occurs, you can assume the equipment used was sterile. However, if
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Introduction to Science microbial growth does occur, you must assume that the equipment was contaminated prior to the experiment and must redo the experiment with new materials.
Alternatively, positive controls are data samples in which you do expect a change. Let’s return to the growth example, but now you need to create a positive control. To do this, you now use a sterile loop to streak a plate with a bacterial sample that you know grows well on agar (such as E. coli). If bacteria grows, you can assume that the bacteria sample and agar are both suitable for the experiment. However, if bacteria does not grow, you must assume that the agar or bacteria has been compromised; the agar is inhibiting growth, or the bacteria in the sample are not viable.
Collecting and Presenting Data
The scientific method also requires data collection. This may reflect what occurred before, during, or after an experiment. Collected data help reveal experimental results. Data should include all relevant observations, both quantitative and qualitative.
After results are collected, they can be analyzed. Data analysis often involves a variety of calculations, conversions, graphs, tables, etc. A common task a scientist faces is unit conversion. Units of time are often displayed in an increment that must be converted. For example, suppose half of your data is measured in seconds, but the other half is measured in minutes. It will be difficult to understand the relationship between the data if the units are not equivalent (sample calculation below).
Significant Digits
When calculating a unit conversion, significant digits must be accounted for. Significant digits are the digits in a number or answer that describe how precise the value actually is. Consider the rules in Table 1.
Addition and subtraction problems should result in an answer that has the same number of significant decimal places as the least precise number in the calculation. Multiplication and division problems should keep the same total number of significant digits as the least precise number in the calculation. For example:
Addition Problem: 12.689 + 5.2 = 17.889
Multiplication Problem: 28.8 x 54.76 = 1577.088
round to 18
round to 1580 (3 significant digits)
Scientific notation is another common method used to report a number. Scientific data is often very large
(e.g., the speed of light) or very small (e.g., the diameter of a cell). Scientific notation provides an abbreviated expression of a number, so that scientists don’t get caught up counting a long series of zeroes.
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Introduction to Science
Table 1: Significant Digits Rules
Rule
Examples
•
45 has two significant digits
•
3.99 has three significant digits
Any time a zero appears between significant numbers, the zero is significant.
•
4005 has four significant digits
•
0.3400000009 has ten significant digits
Zeros that are ending numbers after a decimal point or zeros that are after significant numbers before a decimal point are significant.
•
45.00 has four significant digits
•
15000.00 has seven significant digits
Zeros that are used as placeholders before other digits are NOT significant digits.
•
.0000000897 has three significant digits
A zero at the end of a number with no decimal can be a significant digit.
*To avoid uncertainty, numbers can be written using scientific notation.
•
6200 can have 2, 3, or 4 significant digits
(e.g., 6.2 x 104 has 2, 6.20 x 104 has 3, and
6.200 x 104 has 4)
Any non-zero number (1 - 9) is always significant.
There are three parts to scientific notation: the base, the coefficient and the exponent. Base 10 is almost always used and makes the notation easy to translate. The coefficient is always a number between 1 and 10, and uses the significant digits of the original number. The exponent tells us whether the number is greater or less than 1, and can be used to “count” the number of digits the decimal must be moved to translate the number to regular notation. A negative exponent tells you to move the decimal to the left, while a positive one tells you to move it to the right.
Figure 4: The exponent equals the number of decimal places moved until the co- For example, the number 5,600,000 can be written in scientific noefficient is a number between 1 and 10.
6
tation as 5.6 x 10 . The coefficient is 5.6, the base is 10, and the exponent is 6. If you multiply 5.6 by 10 six times, you will arrive at 5,600,000. Note the exponent, 6, is positive because the number is larger than one. Alternative, the number 0.00045 must be written using a negative exponent. To write this number in scientific notation, determine the coefficient. Remember that the coefficient must be between 1 and 10. The significant digits are 4 and 5. Therefore, 4.5 is the coefficient. To determine the exponent, count how many places you must move the decimal over to create the original number. Moving to the left, we have 0.45, 0.045, 0.0045, and finally 0.00045. Since we move the decimal 4 places to the left, the exponent is -4. Written in scientific notation, we have 4.5 x 10-4
Although these calculations may feel laborious, a well-calculated presentation can transform data into a format that scientists can more easily understand and learn from. Some of the most common methods of data presentation are tables and graphs.
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Introduction to Science
Table: A well-organized summary of data collected. Tables should display any information relevant to the hypothesis. Always include a clearly stated title, labeled columns and rows, and measurement units.
Table 2: Plant Growth With and Without Added Nutrients
Variable
Height Wk. 1 (mm)
Height Wk. 2 (mm)
Height Wk. 3 (mm)
Height Wk. 4 (mm)
Control
3.4
3.6
3.7
4.0
3.5
3.7
4.1
4.6
(without nutrients)
Independent
(with nutrients)
Graph: A visual representation of the relationship between the independent and dependent variable. Graphs are useful in identifying trends and illustrating findings. Rules to remember:
•
The independent variable is always graphed on the x-axis (horizontal), with the dependent variable on the y axis (vertical).
•
Use appropriate numerical spacing when plotting the graph, with the lower numbers starting on both the lower and left hand corners.
•
Always use uniform or logarithmic intervals. For example, if you begin by numbering, 0, 10, 20, do not jump to 25 then to 32.
•
Title the graph and both the x and y axes such that they correspond to the data table from which they come. For example, if you titled your table “Heart rate of those who eat vegetables and those who do not eat vegetables”, the graph title should reference this information as well.
•
Determine the most appropriate type of graph. Typically, line and bar graphs are the most common. Line Graph: Shows the relationship between variables using plotted points that are connected with a line. There must be a direct relationship and dependence between each point connected.
More than one set of data can be presented on a line graph. Figure 5 uses the data from
Table 2.
Plant growth, with and without nutrients, over time.
Figure 5: Sample line graph.
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Introduction to Science
Speed (kph)
Bar Graph: Used to compare results that are independent from each other, as opposed to a continuous series. Since the results from our previous example are continuous, they are not appropriate for a bar graph.
Figure 6: Sample bar graph. Top speed for Cars A, B, C, and D.
After compiling the data, scientists analyze the data to determine if the experiment supports or refutes the hypothesis. If the hypothesis is supported, you may want to consider additional variables that should be examined. If your data does not provide clear results, you may want to consider running additional trials or revising the procedure to create a more precise outcome.
Percent Error
One way to analyze data is to calculate percent error. Many experiments perform trials which calculate known values. When this happens, you can compare experimental results to known values and calculate percent error. Low percent error (<5%) indicates that results are probably accurate, and high percent error
(>20%) indicates that results may be inaccurate. The formula for percent error is:
Percent Error = |(Experimental - Actual)| x 100%
Actual
Note that the brackets flanking the numerator indicate “absolute value”. This means that the number in the equation is always positive.
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Introduction to Science
Suppose your experiment involves gravity. Your experimental results indicate that the speed of gravity is
10.1 m/s2, but the known value for gravity is 9.8 m/s2. We can calculate the percent error through the following steps:
Percent Error = |(10.1 m/s2 - 9.8 m/s2)|
x 100%
(9.8 m/s2)
Percent Error = |0.3|
x 100%
(Note the units cancel each other out)
(9.8)
Percent Error = 0.0306 x 100% = 3.1% (Remember the significant digits)
Writing a Lab Report
The scientific method gives us a great foundation to conduct scientific reasoning. The more data and observations we are able to make, the more we are able to accurately reason through the natural phenomena which occur in our daily lives. Scientific reasoning does not always include a structured lab report, but it always helps society to think through difficult concepts and determine solutions. For example, scientific reasoning can be used to create a response to the changing global climate, develop medical solutions to health concerns, or even learn about subatomic particles and tendencies.
Although the scientific method and scientific reasoning can guide society through critical or abstract thinking, the scientific industry typically promotes lab reports as a universal method of data analysis and presentation. In general terms, a lab report is a scientific paper describing the premise of an experiment, the procedures taken, and the results of the study. They provide a written record of what took place to help others learn and expedite future experimental processes.
Though most lab reports go unpublished, it is important to write a report that accurately characterizes the experiment performed.
Figure 7: Lab reports are an important part of science, providing a way to report conclusions and ideas.
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Introduction to Science
Table 3 summarizes the components of a typical lab report.
Table 3: Lab Report Components
Lab Report
Purpose
Section
Title
Abstract
A short statement summarizing the topic
A brief summary of the methods, results and conclusions. It should not exceed
200 words and should be the last part written.
An overview of why the experiment was conducted. It should include:
Introduction
•
Background - Provide an overview of what is already known and what questions remain unresolved. Be sure the reader is given enough information to know why and how the experiment was performed.
•
Objective - Explain the purpose of the experiment (i.e. "I want to determine if taking baby aspirin every day prevents second heart attacks.")
•
Hypothesis - This is your "guess" as to what will happen when you do the experiment. Materials and
Methods
A detailed description of what was used to conduct the experiment, what was actually done (step by step) and how it was done. The description should be exact enough that someone reading the report can replicate the experiment.
Results
Data and observations obtained during the experiment. This section should be clear and concise. Tables and graphs are often appropriate in this section. Interpretations should not be included here.
Data interpretations and experimental conclusions.
Discussion
32
•
Discuss the meaning of your findings. Look for common themes, relationships and points that perhaps generate more questions.
•
When appropriate, discuss outside factors (i.e. temperature, time of day, etc.) that may have played a role in the experiment.
•
Identify what could be done to control for these factors in future experiments.
Conclusion
A short, concise summary that states what has been learned.
References
Any articles, books, magazines, interviews, newspapers, etc. that were used to support your background, experimental protocols, discussions and conclusions.
Introduction to Science
Exercise 1: Data Interpretation
Dissolved oxygen is oxygen that is trapped in a fluid, such as water. Since many living organism requires oxygen to survive, it is a necessary component of water systems such as streams, lakes and rivers in order to support aquatic life. The dissolved oxygen is measured in units of ppm (parts per million). Examine the data in
Table 4 showing the amount of dissolved oxygen present and the number of fish observed in the body of water the sample was taken from; finally, answer the questions below.
Table 4: Water Quality vs. Fish Population
Dissolved Oxygen (ppm)
0
2
4
6
8
10
12
14
16
18
Number of Fish Observed
0
1
3
10 12
13
15
10
12
13
1. What patterns do you observe based on the information in Table 4?
2. Develop a hypothesis relating to the amount of dissolved oxygen measured in the water sample and the number of fish observed in the body of water.
3. What would your experimental approach be to test this hypothesis?
4. What would be the independent and dependent variables?
5. What would be your control?
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Introduction to Science
6. What type of graph would be appropriate for this data set? Why?
7. Graph the data from Table 4: Water Quality vs. Fish Population (found at the beginning of this exercise).
8. Interpret the data from the graph made in Question 7.
Exercise 2: Testable Observations
Determine which of the following observations are testable. For those that are testable:
Determine if the observation is qualitative or quantitative
Write a hypothesis and null hypothesis
What would be your experimental approach?
What are the dependent and independent variables?
What are your controls - both positive and negative?
How will you collect your data?
How will you present your data (charts, graphs, types)?
How will you analyze your data?
Observations
1. A plant grows three inches faster per day when placed on a window sill than it does when placed on a on a coffee table in the middle of the living room.
2. The teller at the bank with brown hair and brown eyes is taller than the other tellers.
34
Introduction to Science
3. When Sally eats healthy foods and exercises regularly, her blood pressure is 10 points lower than when she does not exercise and eats fatty foods.
4. The Italian restaurant across the street closes at 9 pm but the one two blocks away closes at 10 pm.
5. For the past two days, the clouds have come out at 3 pm and it has started raining at 3:15 pm.
6. George did not sleep at all the night following the start of daylight savings.
Exercise 3: Conversion
For each of the following, convert each value into the designated units. 1. 46,756,790 mg = _______ kg
2. 5.6 hours = ________ seconds
3. 13.5 cm = ________ inches
4. 47 °C = _______ °F
35
Introduction to Science
Exercise 4: Accuracy and Precision
For the following, determine whether the information is accurate, precise, both or neither.
1. During gym class, four students decided to see if they could beat the norm of 45 sit-ups in a minute. The first student did 64 sit-ups, the second did 69, the third did 65, and the fourth did 67.
2. The average score for the 5th grade math test is 89.5. The top 5th graders took the test and scored 89,
93, 91 and 87.
3. Yesterday the temperature was 89 °F, tomorrow it’s supposed to be 88 °F and the next day it’s supposed to be 90 °F, even though the average for September is only 75 °F degrees!
4. Four friends decided to go out and play horseshoes. They took a picture of their results shown to the right:
5. A local grocery store was holding a contest to see who could most closely guess the number of pennies that they had inside a large jar. The first six people guessed the numbers 735, 209, 390, 300, 1005 and
689. The grocery clerk said the jar actually contains 568 pennies.
36
Introduction to Science
Exercise 5: Significant Digits and Scientific Notation
Part 1: Determine the number of significant digits in each number and write out the specific significant digits.
1. 405000
2. 0.0098
3. 39.999999
4. 13.00
5. 80,000,089
6. 55,430.00
7. 0.000033
8. 620.03080
Part 2: Write the numbers below in scientific notation, incorporating what you know about significant digits.
1. 70,000,000,000
2. 0.000000048
3. 67,890,000
4. 70,500
5. 450,900,800
6. 0.009045
7. 0.023
37
Introduction to Science
Exercise 6: Percentage Error
In the questions below, determine the percentage error. Show your work on all problems.
1. A dad holds five coins in his hand. He tells his son that if he can guess the amount of money he is holding within 5% error he can have the money. The son guesses that he is holding 81 cents. The dad opens his hand and displays 90 cents. Did the son guess close enough to receive the money from his father? 2. A science teacher tells her class that their final project requires the students to measure a specific variable and determine the velocity of a car with no more than 2.5% error. Jennifer and Johnny work hard and decide the velocity of the car is 34.87 m/s. The teacher informs them that the actual velocity is 34.15 m/s. Will Jennifer and Johnny pass their final project?
3. A locomotive train is on its way from Chicago, IL to Madison, WI. The trip is said to last 3.15 hours. When the train arrives in Madison the conductor notices it actually took them 3.26 hours. The train company prides itself on always having its trains to the station within a 3% error of the expected time. Will the train company live up to its reputation on this trip?
4. A coach tells his little league players that hitting a 0.275 batting average, within 7% percentage error, means that they had a really great season. Seven year old Tommy ended the season hitting a 0.258 batting average. According to his coach, did he have a great season?
38
Introduction to Science
Exercise 7: Experimental Variables
Determine the variables tested in the each of the following experiments. If applicable, determine and identify any positive or negative controls.
1. A study is being done to test the affects of habitat space on the size of fish populations. Different sized aquariums are set up with six goldfish in each one. Over a period of six months, the fish are fed the same type and amount of food. The aquariums are equally maintained and cleaned throughout the experiment.
The temperature of the water is kept constant. At the end of the experiment the number of surviving fish are surveyed.
A. Independent Variable:
B. Dependent Variable:
C. Controlled Variables/Constants:
D. Experimental Controls/Control Groups:
2. To determine if the type of agar affects bacterial growth, a scientist cultures E. coli on four different types of agar. Five petri dishes are set up to collect results:
•
One with nutrient agar and E. coli
•
One with mannitol-salt agar and E. coli
•
One with MacConkey agar and E. coli
•
One with LB agar and E. coli
•
One with nutrient agar but NO E. coli
All of the petri dishes received the same volume of agar, and were the same shape and size. During the experiment, the temperature at which the petri dishes were stored, and at the air quality remained the same.
After one week the amount of bacterial growth was measured.
A. Independent Variable:
39
Introduction to Science
B. Dependent Variable:
C. Controlled Variables/Constants:
D. Experimental Controls/Control Groups:
40
Lab 2
General Lab Safety
General Lab Safety
Learning Objectives
•
Explain how to safely work in a biological and chemical laboratory
•
Explain how and when to use the safety equipment in a biological and chemical laboratory
•
Identify common laboratory equipment
•
Properly neutralize and dispose of acidic and basic solutions
Introduction
If you’ve ever seen the Frankenstein movies, then you are probably familiar with Hollywood’s interpretation of a laboratory as a dark, dungeonesque space with a mad scientist in action. However, although lab experiences may often appear haphazard in movies, real-world scientists develop and incorporate organized protocols to keep lab environments safe. In addition to focusing on safe lab practices, the experiments in this lab manual will help you to learn about “green”, environmentally-friendly chemicals and adhere to safety precautions.
Before beginning these labs, you must first realize there are specific techniques and precautions to learn to decrease the risks involved. You will need to learn the location of certain items, rules, and the names of common lab equipment. After you get acquainted with your surroundings, you will be prepared and comfortable completing the labs in this manual.
This manual is written for a “Green Chemistry” approach. This means the activities in this manual are generally safer and produce less hazardous waste than similar experiments found in other lab manuals. The chemicals used are environmentally friendly and can be discarded down a household drain with running water. Each of these experiments has been performed safely by other students. However, safer and less hazardous waste does not mean accidents, injuries, or damage will not happen. Scrapes and burns are always a possibility. These experiments will likely be performed in a household environment, so be sure a working fire extinguisher is accessible. You are advised to always use the safety glasses and gloves provided in your kit. Note that the gloves provided in your lab kit are nitrile-based, rather than latex. Please contact eScience
Labs at 1-888-ESL-KITS or info@eScienceLabs.com if you require and alternate.
Safety Equipment
• Safety Glasses: Safety glasses are used to protect your eyes, and should be worn at all times when in the chemistry laboratory, even if you are not currently working with chemicals.
•
Gloves: At times you may need to wear gloves to protect your hands from harmful chemicals or hot objects. The type of glove needed will depend on the application. For example, oven mitts are worn to remove hot objects from an oven while vinyl or latex gloves are used when working with
43
General Lab Safety acids and bases. It is very important not to touch your work area with gloves that have been contaminated with harmful chemicals.
•
Safety Shower: A safety shower is used when a hazardous chemical is spilled on a person where they are unable to rinse it off thoroughly in the sink. It can also be used if a person’s clothes catch on fire. Most safety showers are operated by pulling a chain, although a standard home shower will also work. If a sink is insufficient to thoroughly rinse yourself, get to the nearest shower in your home as quickly as possible. This is not a time for modesty. Remove the contaminated clothing while rinsing your skin with a copious amount of water.
Figure 1: Using the information in this section, you will be able to complete the labs experiments in this manual safely and successfully. Can you identify what’s wrong with this picture?
•
Eye Wash: An eye wash is used if a harmful chemical is splashed into your eyes or face. It is usually operated by pushing forward on a handle. In the home, find the nearest sink and flush water in your eyes for at least 20 minutes. If your sink has a sprayer, use it to rinse your eyes or face while making sure the water drains into the sink.
•
Fire Extinguisher: A fire extinguisher is used to put out small to medium fires.
•
Laboratory Fume Hood: A laboratory fume hood removes harmful gases and fumes sometimes present when doing an experiment. You should always work in a fume hood whenever you are working with corrosive, noxious, or flammable materials. Chemicals used in this kit will not require the use of a fume hood.
Besides knowing where the safety equipment is located and how and when to use it, there are general safety rules you need to follow in the laboratory. Some of the common safety rules are listed below.
Laboratory Safety Rules
44
1.
Always wear safety glasses. Never wear contact lenses.
2.
Never attempt unauthorized experiments.
3.
Always have someone available to help in the event of an accident.
4.
Never have food, drink, chewing gum, or tobacco in the laboratory.
5.
Always keep your work area free of clutter.
6.
Always wear a protective apron and sensible clothing. This means no loose clothing, bare midriffs, or open-toe shoes.
7.
Know the location of and how to use safety equipment in your home. This includes showers, fire
General Lab Safety extinguishers, and sinks.
8.
Always read the experiment entirely before beginning the procedure.
9.
Always wash hands before leaving the lab.
10. Tie back long hair.
11. Never run or play practical jokes in the experiment area.
12. Place broken glass in a protective container, never loosely in a trash can.
Acid-Base Behavior
In addition to following the general safety rules, chemicals need to be handled properly. In particular, two very important classes of compounds called acids and bases require special attention. These compounds are commonly used reagents in the laboratory; therefore, understanding their proper disposal is beneficial.
Physical differences between acids and bases can be detected by the some of the five senses, including taste and touch. Acids have a sour or tart taste and can produce a stinging sensation to broken skin. For example, lemon juice tastes sour. Alternatively, bases have a bitter taste and a slippery texture. Soap and many cleaning products are bases.
Acids and bases are measured on a scale called pH. pH, or potential hydrogen, is calculated using a mathematical equation that accounts for some of Table 1: pH Range of Common
Foods
the chemical differences in acidic and basic compounds. This scale helps
Food
pH Range us quickly determine if a solution is very acidic, a little acidic, neutral
(neither acidic nor basic), a little basic, or very basic. pH values range
Lime
1.8 - 2.0 from less than 1 to 14. A pH of 1 is highly acidic, a pH of 14 is highly basic
(or alkaline), and a pH of 7 is neutral. Table 1 lists the pH of several foods. pH indicators, which change color under a certain pH level, can be used to determine whether a solution is acidic or basic. For example, litmus paper is made by coating a piece of paper with litmus, which changes color at around a pH of 7. Either red or blue litmus paper can be purchased depending on the experimental needs. Blue litmus paper remains blue when dipped in a base, but turns red when dipped in an acid, while red litmus paper stays red when dipped in an acid, but turns blue when in contact with a base.
Acids and bases can react with each other. In this case, the two opposites cancel each other out resulting in a product that is neither an acid nor a
Soft Drinks
2.0 - 4.0
Apple
3.3 - 3.9
Tomato
4.3 - 4.9
Cheese
4.8 - 6.4
Potato
5.6 - 6.0
Drinking
Water
6.5 - 8.0
Tea
7.2
Eggs
7.6 - 8.0
45
General Lab Safety base. This type of reaction is called a neutralization reaction. Neutralization of an acid or base is a technique frequently used for the proper disposal of the compound. The neutralized product can usually be disposed of by flushing it down a sink.
Keep in mind that in a typical laboratory strong acids and bases, such as hydrochloric acid (HCl) and sodium hydroxide (NaOH), are commonly utilized. As mentioned previously, the experiments in this lab manual are designed with a green approach. That means that the acids and bases, in addition to the other chemicals classes, used in this manual are safe for direct disposal.
Some general guidelines for handling chemicals properly are listed below.
Handling Chemicals
46
1.
Always add acids to water, never water to acids.
2.
Never return unused chemicals to the bottles from where they were first obtained.
3.
Dispose of used chemicals in the proper waste containers and/or as instructed.
4.
Always clean the work area, and put away extra equipment when laboratory work is completed.
5.
Never leave anything unattended while it is being heated or is reacting rapidly.
6.
Never carry out a reaction or heat a substance in a closed system.
7.
Always be careful when working with previously heated objects.
8.
Always replace stoppers or lids on bottles containing chemicals.
9.
Weigh chemicals in weigh boats or on paper provided for that purpose. Never weigh chemicals by placing them directly on the scale.
10.
Label all chemicals clearly and completely.
11.
Read labels carefully before using chemicals.
12.
Always lubricate glass tubing or thermometers before inserting them into rubber stoppers.
13.
Material Safety Data Sheets (MSDS) for all chemicals provided can be found on our website at www.eScienceLabs.com/educators/msds. These sheets contain all needed information regarding the danger, safety and disposal of every chemical. Download, print, and review the MSDS for each chemical prior to using that chemical.
General Lab Safety
Even though you follow all of the safety rules, accidents can still happen. This is why it is so important to
know what to do for each type of accident.
How to Respond to Accidents
1. Chemical Spills (on the Bench or Floor): Be sure to clean up the spill immediately. If the spill involves volatile or flammable materials, such as alcohol, make sure ALL flames in the lab area are extinguished and spark-producing equipment is shutdown. In the case of an acid spill, pour baking soda on the acid before cleaning up. In the case of a base spill, pour vinegar on the base before cleaning it up. All other chemicals used in this manual can be cleaned up as you normally would. If you have any questions, check the MSDS.
2. Hazardous Chemical Spills (on a Person): If the spill covers a large area, the typical course of action is to remove all contaminated clothing while the person is under the safety shower. If it is a small area, flush the area immediately with a large amount of water and then wash it with soap. Check the MSDS for the spilled chemical and follow all instructions. Medical assistance may be necessary.
3. Chemicals Spills (in the Eyes): If a harmful chemical is splashed on your face and/or in your eyes, immediate attention is critical. Call for help and get to the nearest sink. If the chemical splashes on your face, and you have glasses on, KEEP the glasses on. Remove the chemical from your face before you remove the glasses. If a chemical gets in your eyes, hold your eyes open in the eyewash for at least 20 minutes. Even though you should not be wearing contact lenses in the lab, if you are, rinse your eyes thoroughly, remove your contacts, and continue to rinse your eyes. A doctor should examine your eyes as soon as possible.
4. Chemical Ingestion: Check the MSDS immediately. Call 911 or “Poison Help” at 1-800-222-1222.
5. Burns: Flush the area with cool running water for 20 minutes. Medical attention may be necessary.
6. Cuts and Wounds: If a chemical gets into the cut or wound, rinse it off immediately with a large amount of water. Avoid contamination, and check the MSDS.
7. Fire: Fires in a laboratory are often contained in pieces of glassware, such as a beaker. You should not move a beaker that has a chemical burning in it. Instead, extinguish the fire simply by covering the mouth of the beaker with a thin curved piece of glass called a watch glass and turning off the source of the flame. A plate or pie pan can also work in place of a watch glass. If the fire is not easily covered, you can use a fire extinguisher. If the fire is too large to extinguish quickly, clear the home and call the fire department immediately.
•
Clothing fires can be extinguished in a safety shower if it is close by. If it is not very close, you will need to STOP, DROP, and ROLL to quickly smother the fire.
47
General Lab Safety
Additional Resources
If you have any doubt as to what you are supposed to be doing and how to do it safely:
•
STOP!
•
Double-check the manual
•
Check the website www.esciencelabs.com
•
Email: Help@esciencelabs.com
•
Call for help 1-888-ESL-KITS (1-888-375-5487)
•
Contact your instructor
If you have ANY questions or concerns regarding a chemical, read the MSDS for that chemical. The MSDS lists the dangers, storage requirements, exposure treatment and disposal instructions for every chemical.
The MSDS for any chemical supplied by eScience Labs, LLC, can be found at www.eScienceLabs.com/ educators/msds. Pre-Lab Questions
1. What should you always wear to protect your eyes when you are in the chemistry laboratory?
2. Should you add acid to water or water to acid?
3. Where should you dispose of broken glass?
What should you do if you spill a chemical on your hand?
48
General Lab Safety
Exercise 1: What Is It?
A chemical laboratory contains special equipment to use while you are performing an experiment. Locate each of the items pictured on the following pages in your lab kit, and place a check mark in the appropriate place when you find it. After you have completed this, sketch a picture and name any additional items that are located in your lab kit, classroom, or home that are likely to be useful for you in completing these labs.
Beaker
50 mL
250 mL
S r S ck
Graduated Cylinder
10 mL
100 mL
Petri Dish
Test Tube
Pipette
49
General Lab Safety
Include your Drawings Here:
50
General Lab Safety
Experiment 1: Neutralization of Acids and Bases
In this experiment, you will learn how to properly neutralize and dispose of acidic and basic solutions.
Materials
5 mL 4.5% Acetic Acid (vinegar), C2H4O2
4 Weigh Boats
(1) 10 mL Graduated Cylinder
*Water
8 Litmus Test Strips (Neutral)
Permanent Marker
*You Must Provide
2 Pipettes
1 g Sodium Bicarbonate (baking soda), NaHCO3
Procedure
1. Use the permanent marker to label three of the weigh boats as A - C.
2. Measure and pour approximately 5 mL of water into weigh boat “A”.
3. Add 0.5 g sodium bicarbonate to weigh boat “B”.
4. Measure and pour approximately 5 mL of water into weigh boat “B”. Gently pipette the solution up and down until the sodium bicarbonate is fully dissolved in the water.
5. Measure and pour 5 mL acetic acid solution to weigh boat “C”.
6. Use the litmus test strips to determine if the substances in weigh boats A - C are acidic or basic. This is accomplished by briefly dipping an unused strip of the litmus paper in each of the weigh boats. Record your color results in Table 2.
7. Pipette 1 mL of the sodium bicarbonate solution from weigh boat “B” into weigh boat “C”. Gently swirl weigh boat “C” to mix.
8. Develop and record a hypothesis regarding the pH of weigh boat “C”. Record this in the Post-Lab Questions section.
9. Test the pH of weigh boat “C” using new litmus paper. Record your result in Table 3.
10. Repeat Step 9 four more times until all the sodium bicarbonate has been added to weigh boat “C”.
51
General Lab Safety
Table 2: Initial Litmus Test Results
Weigh Boat
Chemical Contents
Litmus Results
Additional Observations
A
B
C
Table 3: Neutralization of an Acid
Amount of Base
Litmus Result
1 mL
2 mL
3 mL
4 mL
5 mL
Post-Lab Questions
1. State your hypothesis (developed in Step 8) here. Be sure to include what you think the pH will be, and why. 2. What is a neutralization reaction?
3. When might neutralization reactions be used in a laboratory setting?
4. At what point was the acetic acid in weigh boat “C” neutralized?
5. What do you think would have been the results if a stronger solution of sodium bicarbonate was used?
Would it take more or less to neutralize the acid? What about a weaker concentration of sodium bicarbonate?
52
Lab 3
Chemical Bonding Fundamentals
Molecule Pictured: Arginine
Chemical Bonding Fundamentals
Learning Objectives
•
Relate matter to atoms, protons, electrons, and neutrons
•
Explain how electron configuration affects the interaction of atoms
•
Identify patterns in the periodic table of elements
•
Compare and contrast covalent and ionic bonding
•
Explain how covalent bonding can result in polar and non-polar molecules
Introduction
A myriad of reactions are constantly occurring within each living organism. These reactions allow an organism to reproduce, grow, move, eat, and perform a great many more functions. In order to properly study these biological reactions, a general understanding of chemistry is necessary.
The term matter is used to describe anything that occupies space and has mass. Take a look around, everything you can see and touch is matter (including you). The atomic theory states that all matter is made of atoms. Although physicists and other scientists have developed ways to split atoms to produce nuclear energy, atoms are considered the smallest indivisible unit of all matter. The word atom, in fact, comes from the
Greek word atomos meaning ‘cannot be cut’ or ‘indivisible’.
Atomic Structure
Atoms contain three types of subatomic particles: electrons (negatively charged particles), protons
(positively charged particles) and neutrons (uncharged, neutral particles). Within an atom, the subatomic particles are arranged so that the proton(s) and neutron(s) form a nucleus. The electrons orbit around the nucleus in a very specific manner. This atomic structure is analogous to the manner in which the planets orbit around the sun in our solar system. In this example, the nucleus represents the sun and the electrons represent the orbiting planets.
Instead of gravity holding the atom together, the positive charge of the protons attracts the negative electrons in an electrostatic manner. The configuration of electrons associated with an atom determines the chemical properties of that atom. Having a basic understanding of electron con- Figure 1: Diatomic oxygen molecules share four figuration aids in the overall comprehension of how at- valence electrons through covalent bonding. oms interact. As mentioned above, electrons are found in Note the electron orbitals and the proton-nucleus orbits that surround the nucleus. There are specific orbits bundle located near the center of each atom.
55
Chemical Bonding Fundamentals
(or “shells”) that contain electrons. These shells get filled from the innermost (closest to the nucleus) to the outermost shell. An atom is most stable (or, unreactive) when the outer shell is filled. Some atoms, such as helium, have a filled outer shell. Because of this, those atoms are not reactive. Most atoms do not have filled outer shells. In order to fill their outer shell, atoms can either gain or lose electrons to other atoms, or they can share electrons with other atoms.
Elements are pure substances that are made of only one type of atom. For example, gold is only made up of gold atoms. There are over 100 known elements, each with Figure 2: Cross-section of an atom. different chemical and physical properties. Interestingly, the number of naturally occurring elements is unclear as elements continue to be developed and/or detected in laboratories. The periodic table (Figure 4) has been used to categorize elements.
Figure 3: Elemental information for carbon.
The periodic table contains almost endless useful information for performing biology and chemistry. The periodic table is organized similar to a typical data table. It contains individual cells (compartmentalized squares) that are arranged logically in rows and columns according to specific trends. Each cell contains information about a particular element. In the middle of the cell is a one or two letter abbreviation for a particular element called a chemical symbol. For example, Au is the symbol for gold and Na is the symbol for sodium. Above an element’s symbol is the atomic number, which is the number of protons that exist in an atom of that element. Each element has its own unique number of protons. For example, gold atoms have 79 protons and sodium atoms have 11. The number of electrons is normally equal to the number of protons.
Therefore, the number of electrons in a gold atom is 79 and the number of electrons in sodium atoms is 11.
As you go across a row (called a period) moving left to right in the periodic table, you will find that the atomic number of the elements increases by one. This means the number of protons in the elements increases by one. The addition of that single proton changes the properties of the atom. On the other hand, when moving down a column (called a group) of the periodic table you will find that the number of electrons in the outer shell remains the same. This is why the atoms within a group have similar chemical behavior.
56
Chemical Bonding Fundamentals
Color Key:
Figure 4: The periodic table of elements categorizes all of the known elements. Groups are listed vertically as 1 7. Periods are listed horizontally as 1 - 18.
Chemical Bonding
In nature, most elements are not found alone; atoms of most elements combine to make molecules. A molecule is a mixture of two or more atoms in definite proportions. If the molecule contains atoms of different elements it is called a compound. These atoms are held together by chemical bonds, bringing them to a stable state. Keep in mind that a stable state is accomplished by the filling of the atoms’ outermost shell. Electrons involved with the chemical bonds between atoms are called valence electrons.
The two most common bonds are covalent bonds and ionic bonds. Covalent bonds form when two atoms share valence electrons. Ionic bonds form when an atom or molecule carries an electrical charge, which attracts an atom or molecule of the opposite charge. This electrical charge is created when an atom gains or loses electrons in an attempt to fill its outer shell. The resulting ions have opposite charges and attract one another via electrostatic attractions. Ionic and covalent bonds have very different physical properties. For example, ionic compounds have higher melting and boiling points. Ionic compounds also tend to be electrolytes. This means that they contain free ions which make them electrically conductive.
Ionic compounds are formed when anions (negatively charged chemicals) and cations (positively charged chemicals) bind together with ionic bonds (electrostatic attraction). Sodium chloride, NaCl (table salt), is an
57
Chemical Bonding Fundamentals example of an ionic compound. A sodium ion, having given up an electron (e-) in an attempt to obtain a filled outer shell, is positively charged since the number of protons exceeds the number of electrons.
Na → Na+ + eChloride, on the other hand, accepts an electron to fill its outermost shell.
Cl + e- → ClThe sodium ion (Na+) is the cation and the chloride ion (Cl-) is the anion. A bond can now form between the negatively-charged Cl- and the positively-charged Na+. The above reaction can be written as:
Na+ + Cl- → Na+ClIn covalent bonds, the electrons can be either shared equally or unequally between atoms. When electrons in a covalent bond are share equally, the resulting molecule is non-polar.
There is not overall charge associated with that molecule. Unequal sharing results in a polar molecule. There exists a positive and negative “pole” or area within polar molecules. Fats are typically non-polar while water is a polar molecule. Polar molecules can readily interact with other polar molecules. The positive area of one polar molecule and the negative area of another polar molecule attract each other in an electrostatic manner. What happens when oil is added to water? Do they Figure 5: Molecular polarity is present in wainteract? Polar and non-polar molecules do not, in fact, inter- ter. The negative region is focused near the oxygen molecule, while the positive region is act. focused near the hydrogen molecules.
Figure 6: Olive oil (a fat) is non-polar, while vinegar (a water-based solution) is polar. These two solutions are immiscible, or, cannot form a homogenous mixture.
58
Chemical Bonding Fundamentals
Pre-lab Questions
1. List the atomic numbers for each of the following elements.
Iron
Oxygen
Calcium
Nitrogen
Potassium
Hydrogen
2. What determines if a bond is polar?
3. Use the periodic table to determine if potassium chloride (KCl) formed through covalent or ionic bonds?
Use evidence from the Introduction to support your answer.
4. Research two common, polar molecules and two common nonpolar molecules. Draw their molecular structure and explain how the structure makes each molecule polar or non-polar.
Experiment 1: Slime Time
Inks can be polar or non-polar. Polar solvents pick up polar inks, while non-polar solvents pick up non-polar inks. In this experiment, you will use inks to identify slime and silly putty as polar or non-polar. You will also use paper chromatography to verify the inks are correctly identified as polar or non-polar.
Materials
(1) 250 mL Beaker
Silly Putty®
5 mL 4% Borax Solution, Na2B4O7·10H2O
Dry Erase Marker
Ruler
(1) 10 mL Graduated Cylinder
Uni-ball® Roller Pen
(1) 100 mL Graduated Cylinder
*Distilled or Tap Water
Filter Paper (Disk)
*Newspaper
Filter Paper (Square)
*Notebook Paper
0.5 g Guar Gum
*Scissors
Wooden Stir Stick
Highlighter
Permanent Marker
*You Must Provide
1 Popsicle Stick
59
Chemical Bonding Fundamentals
Procedure
Part 1: Making Slime
1. Weigh out 0.5 g of guar gum into a 250 mL beaker.
2. Measure 50.0 mL of distilled water into a 100 mL graduated cylinder and pour it into the 250 mL beaker that contains the guar gum.
3. Rapidly stir the mixture with a wooden stir stick for three minutes, or until the guar gum is dissolved.
4. Measure 4.00 mL of a 4% Borax solution into a 10 mL graduated cylinder and add it to the guar gum and water. 5. Stir the solution until it becomes slime. This will take a few minutes. If the slime remains too runny, add an additional 1.0 mL of the 4.0% Borax solution and continue to stir until the slime is the slightly runny or gooey. 6. Once you are satisfied with the slime, pour it into your hands. Be sure not to drop any of it on to the floor.
7. Manipulate the slime in your hands. Write down observations made about how slime pours, stretches, breaks, etc. in Part 1 of the Data section.
CAUTION: Slime is slippery and, if dropped, it can make the work area slick.
8. Place the slime back into the beaker and WASH YOUR HANDS.
Part 2: Slime and Putty Ink Tests
1. On a piece of notebook paper make one 20 - 25 mm long mark of each of the inks you are testing
(permanent marker, highlighter, Dry Erase, and Uni-ball® Roller Pen). Space the marks at least one inch apart. Use a pencil to label each mark with its description.
a. Water soluble inks include those in highlighters and certain pens.
b. Water insoluble inks include those in a permanent pen/markers, newsprint, and a dry-erase markers. 2. While the inks are drying, select a passage or a picture in the newspaper to test with the slime.
3. Develop a hypothesis stating whether or not you believe the slime produced in Part 1 will pick up newsprint ink. Record this hypothesis in the Post-Lab Questions section. Then, break off a small piece of slime that is 3 - 5 cm in diameter. Gently place this piece on top of the newspaper print, then carefully pick it up again.
4. Observe and record in Table 1 whether or not the ink was picked up onto the slime.
60
Chemical Bonding Fundamentals
5. Break off another small piece of slime. Once the inks from Step 1 have dried gently place the slime on top of the first spot on the notebook paper, then carefully pick it up. Repeat this for each of the inks. Observe and record which inks were picked up (dissolved) by the slime in Table 1.
6. Repeat this ink testing two more times for accuracy.
7. Hypothesize which inks the silly putty will pick up in the Part 2 of the Data section. Then, perform the ink tests with the Silly Putty® according to the procedure outlined in Steps 5 - 6.
Part 3: Chromatography of Ink Samples
1. Use a pencil or scissors to poke a small hole in the center of a piece of filter paper (see Figure 7).
Folded wick
2. Spot the filter paper evenly spaced approximately 2 cm from the small hole with the two insoluble inks and the
Ink spots
two soluble inks that were used in Part 2, Step 1.
3. Obtain a ½ piece of filter paper. Fold the paper in half several times so that it makes a narrow wick.
4. Insert the wick into the hole of the spotted paper so that it is above the top of the filter paper by approximately 2 cm.
5. Fill a 250 mL beaker 3/4 full with water.
6. Set the filter paper on top of the beaker so that the bottom of the wick is in the water. The paper should hang over the edge of the beaker with the spotted side up.
7. Allow water to travel until it is approximately 1 cm from
Figure 7: Chromatography apparatus for
Procedure Part 3.
the edge of the filter paper. Remove the filter paper from the beaker.
8. Observe which inks moved from where they were originally spotted. Record your observations in Part 3 of the Data section.
61
Chemical Bonding Fundamentals
Table 1: Results of Ink Testing for Silly Putty®
Picked up (dissolved)
Name of Ink
Trial 1
Trial 2
Trial 3
Newsprint
Highlighter
Uni-ball® Roller Pen
Permanent Marker
Dry Erase Marker
Data
Part 1
•
Slime Observations:
Part 2
•
Hypothesis for Silly Putty ® (Procedure Part 2, Step 7):
Part 3
• Observations of inks following chromatography:
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Did not pick up
Trial 1
Trial 2
Trial 3
Chemical Bonding Fundamentals
Post-Lab Questions
1. Record your hypothesis regarding the slime’s ability to pick up newsprint ink here.
2. Did the slime pick up water soluble or water insoluble inks? From these results, what can you conclude about the polarity of slime molecules?
3. Explain how you determined your hypothesis about whether or not silly putty would pick up water soluble inks. Was your hypothesis correct?
4. Were the inks you used properly classified as soluble and insoluble? Explain your answer.
63
Lab 4
Introduction to the Microscope
Introduction to the Microscope
Learning Objectives
•
Demonstrate and explain how to use a microscope
•
Explain how different types of microscopes affect magnification and resolving power
•
Identify parts of the a microscope
Introduction
Microbiology is the study of microorganisms. Microorganisms are organisms that are too small to be seen with the human eye. This is a huge field for such small subjects.
There are many times more microorganisms on Earth than all the other organisms put together. It is estimated, for example, that in the human body alone there are 10 microorganisms for every one human cell. That means that 90% of the cells found in the human body are not human! There are many other objects besides microorganisms, such as biological structures, that cannot be seen with the human eye. What type of objects can you think of that cannot be seen without the aid of some magnification? Biological structures might include a valve found in a vein that prevents blood from moving backwards or the cells that make up a multicellular organism. The fact is Figure 1: Bacterial species exhibit differing shapes. that there are an unimaginable number of objects that we cannot see with the naked eye.
Types of Microscopes
Microscopes are used for the visualization of objects too small for us to see. There are many types of microscopes that range from low-level magnification (e.g., hand-held magnification lens) to mid-level magnification
(e.g., compound light microscopes) to very high-power magnification (e.g., an electron microscope). The type of microscope that is used depends on the application. For example, if you were trying to locate and remove a small splinter from your finger, a hand-help magnification lens (magnifying glass) would be extremely helpful. If you wanted to examine the structure of a tiny virus, you would need an electron microscope.
67
Introduction to the Microscope
Magnification
Magnification is one characteristic to consider when deciding what type of microscope is needed for a particular application. The other key characteristic is the resolving power. The resolving power refers to the ability of a microscope to show detail. Having a device that can greatly magnify an object is helpful only if the magnified object is well defined (not fuzzy). The resolution is largely dependent on the wavelength of light used. Shorter wavelengths produce higher resolutions. Light microscopes use visible light and electron microscopes use a beam of electrons that travel in wavelike patterns. These waves are 100,000 times shorter than visible light waves! Electron microscopy can therefore be used to visualize extremely small structures.
The compound light microscope is the most commonly used type of microscope. The name compound refers to presence of more than one lens. The use of these microscopes can be traced to the late 1500s. The early compound microscopes could magnify objects up to about 10X (this means 10 times). In the 1600s a
Dutch linen merchant, Antonie van Leeuwenhoek, greatly improved upon the production of lens and was able to create microscopes that could magnify objects 200X. He observed and recorded many types of microorganisms with his microscopes. Because of this, van Leeuwenhoek is frequently referred to as “the Father of
Microbiology”. Modern day compound light microscopes can typically magnify objects up to 1000X.
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Figure 2: Microscope components.
Introduction to the Microscope
A representative compound light microscope is shown in Figure 2. Note the two sets of lenses:
•
The ocular lenses (close to your eyes)
•
The objective lenses (close to the “object” on the stage)
Along with a light source, these lenses work together to magnify the object being viewed. In the case of the compound light microscope, the total magnification is equal to the magnification power of the ocular lens multiplied by the magnification power of the objective lens. For example, if the ocular lens magnifies 10X and the objective lens magnifies 10X, the total magnification is 100X.
Microscope Components
Below is a list of the parts of a compound light microscope. Refer to Figure 2 as you read through this list to familiarize yourself with these parts.
•
Base: The flat support of the microscope.
•
Light: Illuminates the object being viewed. This can be either in the form of a light source or a mirror that reflects ambient light onto the image. In the latter case it is important to be working in an environment with adequate ambient light.
•
Stage: Supports the slide or other material to be viewed.
•
Diaphragm: Controls the amount of light allowed on the object.
•
Stage Clips: Secure the slide in place.
•
Revolving Nosepiece: Rotates the objective lenses of different magnifications and allows one of them to be positioned over the slide.
•
Arm: Connects the lower base and the upper head of the microscope (also used to carry the microscope). •
Head: Supports both the ocular lens and the revolving nosepiece.
•
Ocular Lens (eyepiece): The lenses on the microscope typically have a magnification of 10X. If your microscope has a pointer, which is used to indicate a specific area of the specimen, it is attached here. Monocular Microscopes have a single ocular eyepiece while binocular microscopes have two ocular eyepieces.
69
Introduction to the Microscope
How to Use a Microscope
The following steps describe the proper use of a compound light microscope.
1. Always carry a microscope with one hand securely around the arm and the other underneath the base for support.
2. Place the microscope on a table, plug it in, and turn on the light source (or adjust the mirror as necessary). Note: When cleaning a microscope, do not use paper towels or cloths as this will scratch the lens. To preserve the microscope, use only lens paper that will not scratch the optics.
3. To prevent damage to the lens or slides, always start and end with the scanning power objective lens (the shortest one) above the light source.
4. Place your slide on the stage and secure it with the stage clips. It is helpful to visually orient the slide so the object to be viewed is directly in the middle of the opening in the stage where the light is directed up toward the slide.
5. Turn the course adjustment knob to bring the stage all the way up to the scanning power objective lens. While looking through the lens, use the course adjustment knob to slowly lower the stage until the specimen comes into focus.
Note: When using a binocular microscope, adjust the distance between the two oculars until only one object is seen. Record this distance and set your microscope to this distance every time you use it. If someone else uses the microscope, the lenses may be re-adjusted for their eyes.
6. To adjust the light, open or close the diaphragm located over the light source. When properly illuminated, the specimen should not be gray or exceptionally bright.
7. With the object is in general focus, rotate the revolving nosepiece to the low-power lens (the next longest). After focusing with the course adjustment knob, switch to the fine adjustment knob to obtain more precise and greater detail. It may also be necessary to adjust the light, because more light reduces contrast (sharpness).
8. To become familiar with the mechanical stage knobs around the base of the microscope (if present), turn one slowly to the right, noting that the image will be moving toward the left. This image inversion is caused by the lenses.
9. If you need higher magnification, slowly rotate the high-power lens into place (the next longest lens). This will bring the tip of the lenses very close to the slide.
10. Make sure the objective lens does not touch the slide.
11. Whenever you use the high-power lens, only use the fine adjustment knob. If the object was well focused while viewing with the low-power lens, very little adjustment should be necessary.
70
Introduction to the Microscope
12. If you cannot bring the object into focus, return to the low-power lens, focus the object, and then return to the high-power lens.
13. When finished, move the revolving nosepiece to the scanning objective lens position before removing the slide.
Specimen Preparation
Proper specimen preparation helps produce the best visualization possible with any microscope. Glass slides are used to contain the specimen. Live specimens are usually prepared as wet mounts. In wet mount preparation, the specimen is placed on the slide and a much thinner glass cover slip is placed on top of the specimen. Samples are frequently heated to simultaneously kill and fix (secure) the sample to the glass slide.
Fixed samples are then stained to enhance the observable contrast between the cellular features.
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Did You Know...
Different types of stains may be used depending on the desired outcome. For example, if a specific structure is being studied, one might select a dye that stains only that unique structure. Perhaps the most well-known stain is the Gram stain. The Gram stain was developed in
1884 by Hans Christian Gram. Gram staining is often used to categorize bacteria as
Gram positive or Gram negative. The
Gram stain distinguishes between bacteria with thick cell walls (Gram positive Gram negaƟve bacterial populaƟon. Image courtesy of species) from those with thin cell walls the CDC Image Library.
(Gram negative species). Thick cell walls entrap the crystal violet, resulting in purple colored cells. Thin cell walls do not entrap the crystal violet dye. Instead, the safranin stain is accepted and red colored cells result.
How to Prepare a Wet Mount Slide
1. To make a wet mount for a specimen that is not already in liquid, take a clean slide and place the specimen in the center.
2. Add one drop of water.
Note: For cells that are transparent, it may be necessary to add a small drop of stain as opposed to water.
3. Carefully add a coverslip by placing one end down and slowly lowering the other end.
71
Introduction to the Microscope
Note: If the coverslip is added too quickly, large air bubbles may become trapped which can cause difficulty viewing the slide. If this happens, gently remove the coverslip, add another drop of water and try again.
4. Remove excess liquid on the bottom of the slide or around the edges before it is placed on the microscope to avoid damage to the lens. Just touch a tissue to the edge of the coverslip to draw away the water (this is an example of capillary action).
5. If the specimen is already in liquid, place a drop in the middle of the slide and add the coverslip as you did in Step 3.
Pre-Lab Questions
1. Label the following microscope using at least four of the components described within the Introduction.
72
Introduction to the Microscope
Experiment 1: Virtual Magnification Exercise
Materials
“How Big Is It?” demonstration on Student Portal
Note: Review the directions for signing in to the Student Portal at the beginning of this manual if uncertain how to access this information
Procedure
1. Log into your eScience Student Portal account (www.eScienceLabs.com/portal/) and locate the “How Big
Is It” demonstration located near the end of the Introduction to the Microscope section.
2. Load the animation and beginning with the head of a pin, increase the magnification by clicking the arrows below the picture. Note the relative sizes of the objects on the pinhead.
3. Be sure to notice the magnification bar on the lower portion of the demonstration that shows the magnification required to see the objects.
Post-Lab Questions
1. At what magnification do you first notice the ragweed pollen?
2. Which is bigger, a rhinovirus or E. Coli?
3. Based on the magnification, how many of the E. Coli can fit into the same space as the head of a pin?
4. About how many red blood cells could fit across the diameter of a human hair (again, look at the magnification scale)?
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Introduction to the Microscope Experiment 2: Virtual Microscope
Materials
Virtual Microscope on Student Portal
Note: Review the directions for signing in to the Student Portal at the beginning of this manual if uncertain how to access this information.
Procedure
1. Log into your eScience Labs Student Portal account and locate the Virtual Microscope activity located near the end of the Introduction to the Microscope section.
2. Take a tour of the virtual microscope by clicking the “Start Tour” button on the right hand side and learn how to use the different controls to effectively use the simulation.
3. Once you are comfortable using the virtual microscope, turn the light on. Switch views so that you are looking through the ocular lens and can look at the slides.
4. Select the letter e slide on the top right of the page and examine with the 4X objective lens. Increase the magnification until the letter no longer fits in the field of view. Note this magnification.
5. Select the cheek smear slide on the right side of the page and bring the cells within the red circle into focus using the 4X, then 10X objective lenses.
6. View the slide under the 40X and 100X objective lenses, making sure you stay within the red circle.
7. Next, select the onion root tip slide on the right side of the page to view. Start with the 4X objective lens, and bring the cells within the red circle into focus.
8. Switch to the 10X objective lens and readjust the focus so the slide is clear. Continue looking at the slide under higher magnification using the 40X and 100X objective lenses.
9. On the left hand side of the screen, select the “Try This” box. Under measurement, select the m1 box to open an activity that will instruct you how to measure the letter e. (Remember this number as you will have to report it in Question 5!)
Note: You may have to go through the tutorial tour again in order for the “Try This” box to become available. Be sure to work your work through the entire checklist, and then click “exit” button to prompt the “Try This” button to appear again.
74
Introduction to the Microscope Post-Lab Questions
1. After completing the m1 exercise in the “Try this” section, how tall is the letter e?
2. What is the highest objective lens you can use to see the entire letter e?
3. The nuclei (the structure inside a cell that contains DNA) of the cheek cells have been stained using a special dye so that they appear purple. What shape are they?
4. At high magnification, you may notice that not all of the nuclei in the onion root tip slide appear as the shape you described in the question above. What do they look like?
5. What is the first step normally taken when you look through the ocular lenses?
75
Lab 5
The Chemistry of Life
The Chemistry of Life
Learning Objectives
•
Compare and contrast organic and inorganic molecules
•
Relate hydrogen bonding to macromolecules found in living things
•
Compare and contrast the four major organic macromolecules: lipids, carbohydrates, nucleic acids and proteins
Introduction
There are over 100 known elements in the periodic table. Elements are pure substances made of only one type of atom. Interestingly, the number of naturally occurring elements is unclear. More than 90% of all matter is composed of combinations of just four of the approximately 88 naturally occurring elements: oxygen, carbon, hydrogen, and nitrogen. Living organisms, in general, require about 20 elements naturally occurring elements. The periodic table continues to develop as new elements are synthesized in laboratories.
Figure 1: The periodic table of elements categorizes all of the known elements. Groups are listed vertically as 1 7. Periods are listed horizontally as 1 - 18.
79
The Chemistry of Life
Organic Molecules
Although the term “organic” is frequently used to refer to foods or clothing, in biology and chemistry, the terms organic and inorganic are used to categorize molecules by their origin. All organic molecules are of biological origin. In contrast, inorganic molecules are not of derived from living organisms. Since organic molecules are those that derived from living organisms, it follows that they could not be produced without life. The human body is made up of roughly 27% organic molecules and 73% inorganic molecules. Carbohydrates (sugars) and proteins are examples of organic molecules (note that both of these types of molecules are produced by living organisms).
Figure 2: Glucose, fructose, and galactose are all examples of carbohydrates.
Carbon dioxide (CO2), table salt (NaCl) and water (H2O) are all examples of inorganic molecules. Notice that carbon dioxide (CO2) contains carbon but is not an organic molecule. If a molecule contains carbon it may be organic, but this is not the only requirement. Organic molecules are typically identified by the presence of carbon-hydrogen bonds.
Macromolecules
There are many classes of organic compounds, based on the functional groups they contain. In living organisms, the most important organic com- Figure 3: Carbon dioxide is a pounds belong to a classification of molecules called macromolecules. major gas exhaled by humans.
The term “macromolecule” simply means a large molecule. Recall that mol80
The Chemistry of Life ecules are substances that contain two or more atoms bonded together. The same four types of macromolecules are used by all living organisms for cellular metabolism and reproduction. These common biological macromolecules are lipids, nucleic acids, carbohydrates and protein. The properties they convey are of great importance to cell function.
Lipids, or fats, have many functions within living organisms including energy storage, membrane structure, and aids in the formation of internal cellular components. All genetic material is composed of nucleic acids (DNA). Another nucleic acid, RNA, functions in the production of proteins and other cellular processes. Carbohydrates are commonly referred to as sugars. This class of organic compounds serves as one of the primary sources of metabolic energy.
The carbohydrate monosaccharide subunit most commonly used for energy is glucose. Other useful carbohydrates include maltose, lactose, sucrose, and starch. Proteins are the most abundant macromolecule in living systems. Like lipids, proteins perform a variety of functions. For example, proteins are major
Figure 4: Omega-3 Fatty acid. components of tendons, ligaments and muscles in the human body.
Chemical Bonding
Macromolecules are formed by the covalent bonding between subunits. Recall that covalent bonds can produce molecules that are either polar or non-polar. Lipids, for the most part, are non-polar. Some of the 20 amino acids found in proteins are non-polar and some are polar. When macromolecules are created, non-polar regions tend to be located close together. Polar regions can participate in another type of chemical bond called hydrogen bonding. Hydrogen bonds result from the interaction of the positive region of one polar molecule and the negative region of another polar molecule.
Although relatively weak bonds, these are extremely imFigure 5: Approximately 70% of a human adult portant in the maintenance of chemical structures. In fact, the body is composed of water. double strands found in DNA as well water molecules are held together by hydrogen bonds. Adenine and thymine
(nucleic acids found in DNA) are held together via two hydrogen bonds, and cytosine and guanine are held together by three.
Living things require a constant supply of energy. Throughout this manual, you will learn about the reactions that take place inside of organisms. The sum of these reactions is called metabolism, and is a general term used to describe the energy required to keep those reactions occurring.
81
The Chemistry of Life
Pre-Lab Questions
1.
Nitrogen fixation is a natural process by which inert or unreactive forms of nitrogen are transformed into usable nitrogen. Why is this process important to life?
2. Given what you have learned about the hydrogen bonding shared between nucleic acids in DNA, which pair is more stable under increasing heat: adenine and thymine, or cytosine and guanine? Explain why.
3. Which of the following is not an organic molecule; methane (CH4), fructose (C6H12O6), rosane (C20H36), or ammonia (NH3)? How do you know?
Experiment 1: Testing for Proteins
The protein molecules in many foods provide the amino acid building blocks required by our own cells to produce new proteins. To determine whether a sample contains protein, a reagent called Biuret solution is used. Biuret solution contains copper ions. However, the chemical state of the copper ions in Biuret solution causes them to form a chemical complex with the peptide bonds between amino acids (when present), changing the color of the solution. Biuret solution is normally blue, but changes to pink when short peptides are present and to violet when long polypeptides are present.
82
Figure 6: Biuret solution only is located on the far left side of the image (blue).
Note the transition from blue to violet as proteins are added to the solution, causing the solution to transition from blue to violet.
The Chemistry of Life
Materials
(2) 250 mL Beakers
5 Test Tubes (Plastic)
25 Drops Biuret Solution, H2NC(O)NHC(O)NH
Test Tube Rack
(1) Knox® Gelatin Packet
5 mL Unknown Solution
5 mL 1% Glucose Solution, C6H12O6
*Tap Water
(1) 10 mL Graduated Cylinder
*Hot Water
(1) 100 mL Graduated Cylinder
*Egg White
Permanent Marker
5 Pipettes
*You Must Provide
Procedure
1. Label five test tubes 1, 2, 3, 4 and 5.
2. Prepare your testing samples as follows:
a. Mix one egg white with 25 mL water in a 250 mL beaker to create an albumin solution. Pipette
5 mL of this solution into Test Tube 1.
b. Mix the packet of Knox® gelatin with 50 mL hot water in a second 250 mL beaker. Stir until dissolved. Pipette 5 mL of this solution into Test Tube 2.
3. Pipette 5 mL of the 1% glucose solution into Test Tube 3.
4. Use the 10 mL graduated cylinder to measure and pour 5 mL of water into Test Tube 4.
5. Pipette 5 mL of the “Unknown Solution” into Test Tube 5.
6. Record the initial color of each sample in Table 1.
7. Develop a hypothesis regarding what you predict will happen when Biuret solution is added to Tubes 1
- 4. Record your hypothesis in the Post-Lab Question section. Then, pipette five drops of Biuret solution to each test tube (1 - 5). Swirl each tube to mix.
8. Record the final color in Table 1.
Note: Protein is present in the sample if a light purple color is observed.
83
The Chemistry of Life
Table 1: Testing for Proteins Results
Sample
Initial Color
Final Color
Protein Present
1 - Albumin Solution
2 - Gelatin Solution
3 - Glucose
4 - Water
5 - Unknown
Post-Lab Questions
1. Record your hypothesis about what will happen when Biuret solution is mixed with the solutions from test tubes 1, 2, 3, and 4 here. Be sure to use scientific reasoning to support your hypothesis.
2. Write a statement to explain the molecular composition of the unknown solution based on the results obtained during testing with the Biuret solution and each sample solution.
3. Diet and nutrition are closely linked to the study of biomolecules. How should you monitor your food intake to insure the cells in your body have the materials necessary to function?
84
The Chemistry of Life
4. There are other types of reagents used to determine what type of biomolecule a substance is. For example, copper ions present in Benedict’s reagent reacts with the free end of any reducing sugars, such as glucose, when heated. Originally blue in color, these copper ions are reduced by the sugar, and produce an orange-red colored precipitate. Alternatively, iodine-potassium iodide (IKI) may also be used when working with starch. IKI contains special tri-iodine ions which interact with the coiled structure of a starch polymer. Prior to a reaction, the IKI displays a yellow-brown color; however, after reacting with starch, a dark purple or black color is presented.
The molecule pictured below produced a blue color when tested with Benedict’s reagent, a yellow color when tested with IKI, and a violet color when tested with Biuret reagent. Based on the structure shown below and these chemical results, what kind of biomolecule is this?
85
Lab 6
Diffusion
Diffusion
Learning Objectives
•
Apply diffusion to cellular activity and dialysis
•
Explain how solute size, molecular polarity and membrane permeability affect the rate of diffusion
•
Explain how concentration gradients and molarity affect the direction of diffusion
Introduction
Molecules are constantly in motion due to the kinetic energy present in every atom. This energy results in the net movement of molecules from areas of high concentration to areas of low concentration, a process called diffusion. If uninhibited, this net movement will continue until equilibrium is reached and the molecules are uniformly distributed. Once equilibrium is achieved, molecules will continue to move in each direction at an equal rate.
Diffusion
A solution contains two or more substances (solutes) that have been dissolved by a solvent. In the context of a cell, the intracellular and extracellular fluids are the solvents which contain dissolved material (solutes).
The rate of diffusion depends on membrane characteristics, size of the solute, and molecular polarity. Because the medium will not change in a biological system, the diffusion rate is usually dictated by molecular characteristics of the solute. Small, non-polar molecules exhibit a higher rate of diffusion than large, charged ones.
Diffusion Factors
The direction of diffusion depends on concentration gradients, heat, and pressure. The concentration gradient is the change of molecular density over a given area (density is defined as unit mass per volume of space). Molarity (moles/ liter) is a way to express concentration. It is the number of moles of solute dissolved in one liter of solution, and is commonly represented by ‘M’. A mole equals 6.02 x 1023 molecules or atoms. Molecular weight (MW) is the weight of one mole of a chemical and is based off the atomic mass of each atom in the chemical formula. Once the molecular weight of a chemical is known, the weight of chemical to dissolve in a solvent for a specific molar solution can easily be calculated.
Figure 1: Molecules diffuse down concentra-
Temperature and pressure typically remain constant in biologtion gradients to create equilibrium. ical systems, making the concentration gradient the best indi-
89
Diffusion cator of directionality. In general, molecules will move towards areas of lower concentration.
Diffusion Across a Membrane
A major determinant of diffusion within a biological system is membrane permeability. Cells, as well as some organelles within the cell, are surrounded by selective and differentially permeable membranes. These membranes control the interaction of the cell and its surrounding environment. Acting as a living gatekeeper, the membrane allows, slows, or denies access into the cell.
Cellular membranes are composed of two layers of phospholipids (Figure 1). Each phospholipid has a hydrophobic, fatty acid tail and a hydrophilic head. This lipid bilayer selects for molecules that can dissolve into the lipid environment and against those that cannot. The ability of a molecule to cross the membrane is also determined by its size and electrical charge. Figure 2: Diffusion through a phospholipid
Small, uncharged molecules often pass through the membrane bilayer (cell membrane). easily, while most large or charged molecules are prevented from passing. Molecules which cannot diffuse across the membrane may be able to cross through other regulated gateways located within the membrane.
Dialysis is the separation of molecules through diffusion. In dialysis, a differentially permeable membrane is used to separate the components of a mixed solution containing more than one type of molecule. This membrane allows the free passage of water, but limits the movement of molecules by their size. In one of the following experiments, you will dialyze a solution of glucose and starch to observe membrane permeability.
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Did You Know...
Serial Dilutions
The concentration of a solution may also impact the membrane permeability. A solution or its components may not be able to pass through a membrane if the concentration is too high. For this reason, serial dilutions may be performed to create solutions of lower concentrations. The original sample is a known volume and concentration. A small volume of this sample is transferred sequentially among new tubes, called dilution blanks. The sample is mixed with water (or another diluent) throughout the process to create more dilute solutions (Figure 3).
90
Hemodialysis is a method of removing toxic substances from the blood when the kidneys are unable to do so. It is frequently used for patients with kidney failure, but may also be used to quickly remove drugs or poisons in dangerous situations.
Diffusion
9 mL
9 mL
9 mL
9 mL
9 mL
9 mL
Figure 3: Serial dilution schematic. The dilution blanks all have 9 mL of diluent in them such that
1
when 1 mL is added, it represents /10th the concentration of the previous tube. The dilution of the sample is written below each tube.
Pre-Lab Questions
1. A concentration gradient affects the direction that solutes diffuse. Describe how molecules move with respect to the concentration.
2. How does the size of a solute affect the rate of diffusion? Consider the size and shape of a molecule in your response.
3. Does polarity affect diffusion? Explain your answer using scientific principles.
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Diffusion
Experiment 1: Diffusion through a Liquid
In this experiment, you will observe the effect that different molecular weights have on the ability of dye to travel through a viscous medium.
Materials
1 60 mL Corn Syrup Bottle, C12H22O11
Ruler
Red and Blue Dye Solutions (Blue molecular weight = 793 g/mole; Red molecular weight = 496 g/mole) *Stopwatch
(1) 9 cm Petri Dish (top & bottom halves)
*Tape
*You Must Provide
Procedure
1. Use clear tape to secure one half (either the bottom or the top half is fine) of the petri dish over a ruler.
Make sure that you can read the measurement markings on the ruler through the petri dish. The dish should be positioned with the open end of the dish facing upwards.
2. Carefully fill the half of the petri dish with corn syrup until the entire surface is covered.
3. Develop a hypothesis discussing which dye you believe will diffuse faster across the corn syrup and why.
Record this in the Post-Lab Questions section. Then, place a single drop of blue dye in the middle of the corn syrup. Note the position where the dye fell by reading the location of the outside edge of the dye on ruler. 4. Record the location outside edge of the dye (the distance it has traveled) every ten seconds for a total of two minutes. Record your data in Tables 1 and 2.
5. Repeat Steps 1 - 4 using the red dye, the second half of the petri dish, and fresh corn syrup.
Table 1: Rate of Diffusion in Corn Syrup
Time (sec)
92
Blue Dye
Red Dye
Time (sec)
10
70
20
80
30
90
40
100
50
110
60
120
Blue Dye
Red Dye
Diffusion
Table 2: Speed of Diffusion of Different Molecular Weight Dyes
Structure
Molecular Weight
Total Distance
Traveled (mm)
Speed of Diffusion
(mm/hr)*
Blue Dye
Red Dye
*Multiply the total distance diffused by 30 to get the hourly diffusion rate
Post-Lab Questions
1. Record your hypothesis from Step 3 here. Be sure to validate your predictions with scientific reasoning.
2. Which dye diffused the fastest?
3. Does the rate of diffusion correspond with the molecular weight of the dye?
4. Does the rate of diffusion change over time? Why or why not?
5. Examine the graph below. Does it match the data you recorded in Table 2? Explain why, or why not.
Submit your own plot if necessary.
Diffusion (mm)
Speed of Diffusion of Different Molecular Weight Dyes
Time (seconds)
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Diffusion
Experiment 2: Concentration Gradients and Membrane Permeability
In this experiment, you will dialyze a solution of glucose and starch to observe:
•
The directional movement of glucose and starch.
•
The effect of a selectively permeable membrane on the diffusion of these molecules.
An indicator is a substance that changes color when in the presence of the substance it indicates. In this experiment, IKI will be used an indicator to test for the presence of starch and glucose.
Materials
(5) 100 mL Beakers
*Stopwatch
10 mL 1% Glucose Solution, C6H12O6
*Water
4 Glucose Test Strips
*Scissors
(1) 100 mL Graduated Cylinder
*15.0 cm Dialysis Tubing
4 mL 1% Iodine-Potassium Iodide, IKI
5 mL Liquid Starch, C6H10O5
*You Must Provide
3 Pipettes
*Be sure to measure and cut only the length you
4 Rubber Bands (Small; contain latex, handle with
for later experiments.
gloves on if allergic)
Attention!
Do not allow the open end of the dialysis tubing to fall into the beaker. If it does, remove the tube and rinse thoroughly with water before refilling with a starch/glucose solution and replacing it in the beaker.
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need for this experiment. Reserve the remainder
Note:
•
Dialysis tubing can be rinsed and used again if you make a mistake. •
Dialysis tubing must be soaked in water before you will be able to open it up to create the dialysis “bag”. Follow the directions for the experiment, beginning with soaking the tubing in a beaker of water. Then, place the dialysis tubing between your thumb and forefinger and rub the two digits together in a shearing manner. This should open up the
"tube" so you can fill it with the different solutions.
Diffusion
Procedure
1. Measure and pour 50 mL of water into a 100 mL beaker. Cut a piece of dialysis tubing 15.0 cm long.
Submerge the dialysis tubing in the water for at least 10 minutes.
2. Measure and pour 82 mL water into a second 100 mL beaker. This is the beaker you will put the filled dialysis bag into in Step 9.
3. While the dialysis bag is still soaking, make the glucose/sucrose mixture. Use a graduated pipette to add five mL of glucose solution to a third beaker and label it “Dialysis bag solution”. Use a different graduated pipette to add five mL of starch solution to the same beaker. Mix by pipetting the solution up and down the pipette six times.
4. Using the same pipette that you used to mix the dialysis bag solution, remove two mL of that solution and place it in a clean beaker. This sample will serve as your positive control for glucose and starch.
a. Dip one of the glucose test strips into the two mL of glucose/starch solution in the third beaker.
After one minute has passed, record the final color of the glucose test strip in Table 3. This is your positive control for glucose.
b. Use a pipette to transfer approximately 0.5 mL of IKI to into the two mL of glucose/starch solution in the third beaker. After one minute has passed, record the final color of the glucose/starch solution in the beaker in Table 3. This is your positive control for starch.
5. Using a clean pipette, remove two mL of water from the 82 mL of water you placed in a beaker in Step 2 and place it in a clean beaker. This sample will serve as your negative control for glucose and starch.
a. Dip one of the glucose test strips into the two mL of water in the beaker. After one minute has passed, record the final color of the glucose test strip in Table 3. This is your negative control for glucose. b. Use a pipette to transfer approximately 0.5 mL of IKI to into the two mL of water in the beaker.
After one minute has passed, record the final color of the water in the beaker in Table 3. This is your negative control for starch.
Note: The color results of these controls determine the indicator reagent key. You must use these results to interpret the rest of your results.
6. After at least 10 minutes have passed, remove the dialysis tube and close one end by folding over 3.0 cm of one end (bottom). Fold it again and secure with a rubber band (use two rubber bands if necessary).
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Diffusion
7. Make sure the closed end will not allow a solution to leak out. You can test this by drying off the outside of the dialysis bag with a cloth or paper towel, adding a small amount of water to the bag, and examining the rubber band seal for leakage. Be sure to remove the water from the inside of the bag before continuing.
8. Using the same pipette which was used to mix the solution in Step 3, transfer eight mL of the solution from the Dialysis Bag Solution beaker to the prepared dialysis bag.
9. Place the filled dialysis tube in beaker filled with 80 mL of water with the open end draped over the edge of the beaker as shown in Figure 4.
10. Allow the solution to sit for 60 minutes. Clean and dry all materials except the beaker with the dialysis bag.
11. After the solution has diffused for 60 minutes, remove the dialysis tube from the beaker and empty the contents into a clean, dry beaker. Label it dialysis bag solution.
Figure 4: Step 9 reference.
12. Test the dialysis bag solution for the presence of glucose and starch. Test for the presence of glucose by dipping one glucose test strip into the dialysis bag directly. Again, wait one minute before reading the results of the test strips. Record your results for the presence of glucose and starch in Table 4. Test for the presence of starch by adding two mL IKI. Record the final color in Table 4 after one minute has passed.
13. Test the solution in the beaker for glucose and starch. Use a pipette to transfer eight mL of the solution in the beaker to a clean beaker. Test for the presence of glucose by dipping one glucose test strip into the beaker. Wait one minute before reading the results of the test strip and record the results in Table 4. Add two mL of IKI to the beaker water and record the final color of the beaker solution in Table 4.
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Diffusion
Table 3: Indicator Reagent Data
Indicator
Starch Positive
Starch Negative
Glucose Positive
Glucose Negative
Control (Color)
Control (Color)
Control (Color)
Control (Color)
n/a
n/a
IKI Solution
Glucose Test Strip
n/a
n/a
Table 4: Diffusion of Starch and Glucose Over Time
Indicator
Dialysis Bag After 1 Hour
Beaker Water After 1 Hour
IKI Solution
Glucose Test Strip
Post-Lab Questions
1. Why is it necessary to have positive and negative controls in this experiment?
2. Draw a diagram of the experimental set-up. Use arrows to depict the movement of each substance in the dialysis bag and the beaker.
3. Which substance(s) crossed the dialysis membrane? Support your response with data-based evidence.
4. Which molecules remained inside of the dialysis bag?
5. Did all of the molecules diffuse out of the bag into the beaker? Why or why not?
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Lab 7
Osmosis
Osmosis
Learning Objectives
•
Explain how tonicity affects the direction of osmosis
•
Hypothesize why aquaporins are necessary to support osmosis in living things
•
Relate osmotic pressure to water potential
Introduction
The movement of water across a selectively permeable membrane, like the plasma membrane of the cell, is called osmosis. Osmosis is directed from an area of high water concentration to an area of low water concentration. Ultimately, membrane selectivity and the movement of water in and out of the cell regulate the concentration of intracellular material. As solute concentration increases, solvent concentration decreases.
In order to achieve equilibrium, the solvent from a low solute concentration solution will move into an area of high solute concentration to create an equal ratio of solvent to solute in both areas. Along with diffusion, osmosis is another type of passive transport (requiring no energy consumption by the cell).
Tonicity is a relative term that describes the solute difference between solutions and determines the net direction of movement of water molecules (osmosis). There are three types of tonicity:
1. Hypertonic: A solution with a higher solute concentration than the solute concentration on the opposite side of the permeable membrane.
2. Hypotonic: A solution with a lower solute concentration than the solute concentration (i.e., a solution with a higher percentage of water than solute) on the opposite side of the permeable membrane.
3. Isotonic: A solution with equal solute concentrations on both sides of the permeable membrane
(Figure 1).
Figure 1: Chemical diffusion can create a variety of states for the cell or even an entire organ.
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Osmosis
Water Potential
In osmosis, water molecules flow from hypotonic solutions to hypertonic solutions, until the solutions become isotonic. This underscores the concept of water potential. Biologists use this term to describe the tendency of water to leave one place in favor of another. Water always moves from an area of higher water potential to an area of lower water potential. The more negative the water potential, the higher the concentration of solutes in the system.
In most biological systems, cells are hypertonic to their environment and extracellular water flows into them.
If placed in pure water, cells will burst (lyse) as a result of the increased pressure on the membrane from the additional water that diffused into the cell.
Osmotic Pressure
Osmotic pressure (the force required to prevent osmosis), also called pressure potential, is the tendency of a solution to gain water across an ideal, partially permeable membrane. It directly correlates with tonicity
(higher tonicity causes an increase in osmotic pressure). Some cells, such as plant cells, have specialized structures that regulate osmotic pressure and prevent lysis. This pressure, as well as solute concentration, can affect the water potential (high osmotic pressure = low water potential). This relationship is demonstrated by the following formula:
Ψ = Ψp + Ψs
In this equation, Ψ represents water potential, Ψp is pressure potential, and Ψs is the solute potential.
Both diffusion and osmosis are imperative to cell survival. They enable cells to maintain equilibrium of solute concentrations across two sides of a permeable membrane. However, the two processes facilitate solute equilibrium in opposite ways.
Aquaporins
For much of scientific history, the abundance of water molecules throughout the body led many scientists to believe that osmosis alone was a sufficient mechanism for water movement across cellular membranes.
However, in 1992 Peter Agre discovered a unique family of proteins, later termed “aquaporins”, which were also involved in water transport.
Aquaporins are a specific form of integral membrane proteins which span lipid bilayers to facilitate molecular movement. They use a combination of electromagnetic and hydrophilic interactions and spatial selectivity to funnel water molecules from one side of a membrane to another. Aquaporins are most populous in tissues
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Osmosis which require high levels of water transport such as the kidney, salivary glands, (lacrimal) tear glands and other epithelial cells. Can you think of a reason why aquaporins are so important in these tissues? Although aquaporins can be ubiquitous in certain locations, the actual aquaporin present may vary by tissue/organ.
The most common aquaporin in called aquaporin 1 (AQP1). AQP1 is found in the kidney and forms a homotetramer protein across the cellular membranes within this organ. “Homotetramer” means that it is composed of four, identical simpler molecules.
Water molecules are attracted to these pores. However, rather than moving through the central homotetramer central channel, water molecules move through one of the four individual channels formed by the four monomer subunits. These channels are very tiny, with a diameter of only 3 Å. This further increases the specific of aquaporins because many molecules are much larger than this (note: water molecules are only 2.8
Å).
Agre won the Nobel Prize in Chemistry in 2003 for this paradigm-changing discovery. You can learn more about Peter Agre and his discovery here: http://www.nobelprize.org/nobel_prizes/chemistry/laureates/2003/ agre-autobio.html Pre-Lab Questions
1. What is the water potential of an open beaker containing pure water?
2. Why don’t red blood cells swell or shrink in blood?
3. How do osmotic power plants work?
4. Research the structures that protect plant and animal cells from damage resulting from osmotic pressure.
Write a few paragraphs explaining what they are, how they work, and where they are located.
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Osmosis
Experiment 1: Tonicity and the Animal Cell
In this experiment you will explore the effects of osmosis on eggs. An egg is surrounded by a permeable membrane and a hard shell. To observe the effects of osmosis the shell will first need to be removed.
Materials
Acetic Acid (Vinegar)
100 mL Graduated Cylinder
4 Pieces of Aluminum Foil (to cover beakers)
100% Sugar Solution (Corn Syrup)
(4) 100 mL Beakers
Stirring Rod
*Distilled Water
*4 Eggs
*You Must Provide
10 mL Graduated Cylinder
Note: This lab requires at least one day to prep and at least one day for observations. For best results, please use a medium or large sized egg. Jumbo or extra large eggs are too large for the experimental set-up.
Procedure
1. Set out four beakers with one egg in each beaker.
Note: Always wear gloves when handling raw eggs.
2. Cover each egg with acetic acid (vinegar).
3. Cover each beaker with a piece of foil and allow the eggs to sit for 24 hours.
Keep foil for entire experiment. You will need it later in the experiment.
4. You will be placing eggs (cells) in different tonicities and observing the results.
Develop a hypothesis and record it in the data section for this experiment.
5. After 24 hours, check to see if the egg shell is dissolved by gently taking out
Figure 2: An egg that h as each egg and rinsing with a small amount of water. The eggs should have a been soaked in vinegar and rinsed with water. yellow tint to them and you should be able to lightly squish the egg between
Some white shell remains.
your fingers (Figure 2). BE CAREFUL NOT TO BREAK THE EGG.
6. If any of the eggs still have a hard shell after 24 hours allow the eggs to sit for another 12 - 24 hours.
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Osmosis
7. Once the egg shells have been dissolved remove them from the vinegar. HANDLE EGGS CAREFULLY. THEY CAN EASILY BREAK.
8. Rinse out and dry all four beakers.
9. Label one beaker “100 % distilled water”, one “100% corn syrup solution”, one “10% corn syrup/90% water solution” and one “control”.
10. In the beaker labeled “10% corn syrup/90% water solution”, prepare a 10% sugar solution by mixing 45 mL of distilled water with 5 mL of corn syrup. Mix well with a stirring rod. Record the initial volume in Table 1.
11. In the beaker labeled “distilled water”, measure and pour 50 mL of distilled water. Record the volume in
Table 1. The egg should be covered. Add or reduce the amount of water if necessary. Record the initial volume in Table 1.
12. In the beaker labeled “100% corn syrup solution”, measure and pour 50 mL of corn syrup (100% sugar solution). The egg should be covered. Add more corn syrup if necessary. Record the initial volume of the solution in Table 1.
13. In the beaker labeled “control”, do not add any solution. Record the volume in Table 1.
14. Add one egg to each beaker and cover with foil.
15. Let the eggs sit for 24 hours. In the meantime, develop a hypothesis stating how you believe each egg will be affected by the solution in the beaker over the 24 hour period. Record your hypothesis in the
Post-Lab Questions section.
16. After 24 hours have passed, observe the eggs. If you do not see any noticeable difference let the eggs sit for another 12 - 24 hours.
17. After the eggs have rested for 24 - 48 hours in the solutions carefully remove them from the beakers.
Make sure to note which eggs came from each beaker. Be careful to not spill the solutions.
18. Record the final volume of each solution using a graduated cylinder and record your results in Table 1.
19. Observe the eggs. Draw your observations in the in Table 2 of data section.
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Osmosis
Data
Hypothesis:
Table 1: Osmosis Results
Solution Type
Volume (mL) Before Osmosis
Volume (mL) After Osmosis
100% distilled water
100% corn syrup
10% corn syrup/ 90% distilled water control Table 2: Observations of Eggs After Osmosis
100% Distilled Water
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100% Corn Syrup
10% Corn syrup/ 90%
Distilled Water
Control
Osmosis
Post-Lab Questions
1. Record your hypothesis from Step 15 here. Be sure to include scientific reasoning to support your predictions.
2. How do each of the three eggs placed in solution compare to the control egg?
3. For each beaker, identify whether the solution inside was hypotonic, hypertonic, or isotonic in comparison to the control beaker.
4. What was the direction of osmosis in the beaker labeled “100% distilled water”? Did the egg in this beaker burst? 5. Was there a volume in the “control” beaker after 24-48 hours? If so, why do you think this is? If not, why do you think this is?
6. Which solution contained a dehydrated cell? Think about someone having a high sugar diet. Hypothesize how their cells might be affected by osmosis?
7. Osmosis is how excess salts that accumulate in cells are transferred to the blood stream so they can be removed from the body. Explain how this process works in terms of tonicity.
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Osmosis
Experiment 2: Tonicity and the Plant Cell
Plant cells are able to generate osmotic pressure while other cells cannot. This is due to specialized plant structures such as the cell wall which prevent lysis caused by osmosis. By taking advantage of this system, you will be able to look at the effects of tonicity in a biological system.
Materials
(1) 100 mL Graduated Cylinder
*Kitchen Knife
Permanent Marker
*2 Potatoes (these must be different types; e.g.,
2 Pipettes
Ruler
16 g Sodium Chloride (Salt), NaCl
russet, Idaho, sweet, etc.)
*Stopwatch
*Water
4 Test Tubes (Glass)
Test Tube Rack
*You Must Provide
*Cutting Board
Procedure
1. Use the permanent marker to label two test tubes as A, and two test tubes as B. Place the test tubes in the test tube rack.
2. Identify the two potato types in the first two cells of the first column in Table 3. Then, select one potato to test first and record observations about the physical characteristics in Table 3.
Note: Be sure to include observations which acknowledge the texture, color, and any other distinguishing factors.
3. Use a knife to carefully cut two strips of the potato on a cutting board. These will be referred to as Sample A and Sample B. The strips should be as close to 10.0 cm. long and 1.0 cm. wide as possible to ensure that the strips fit in the test tubes.
4. Fill the 100 mL graduated cylinder with 50 mL of water. Place the first potato strip (Sample A) into the graduated cylinder and record the initial displacement in Table 3.
Note: Displacement is a measurement of change. It is calculated by subtracting the original volume
(50 mL) from the final volume after the potato is added to the 50 mL of water. For example, 57 mL 50 mL = 7 mL of displacement.
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Osmosis
5. Remove Sample A from the graduated cylinder and place it in Test Tube A. If any water was lost in the graduated cylinder, refill it to the 50 mL graduation mark.
6. Place the second potato strip (Sample B) into the graduated cylinder. Record the initial displacement in
Table 3.
7. Remove Sample B from the graduated cylinder and place it in Test Tube B. If any water was lost in the graduated cylinder, refill it to the 50 mL graduation mark.
8. Repeat Steps 2 - 7 for the second potato type, using the remaining test tubes in the test tube rack.
9. Use a pipette to add water to each of the test tubes with the A samples in them until the water covers the potato strips.
10. Refer to the instructions provided on the bottle with 16 g of sodium chloride in it to create a 20% sodium chloride solution. Use a pipette to add the 20% sodium chloride solution to each of the test tubes with the
B potato samples in them until the solution covers the potato strips.
11. After an hour, pour out the liquid from the test tubes.
12. Repeat Steps 4 - 7 for each sample and type, and record the final displacement values in Table 3.
13. Complete the last column of Table 3 by subtracting the initial displacement from the final displacement.
Table 3: Water Displacement per Potato Sample
Potato
Type
Potato
Sample
Observations
Initial Displacement
(mL)
Final Displacement
(mL) - Step 12
Net Displacement
(mL)
A
B
A
B
Post-Lab Questions
1. How did the physical characteristics of the potato vary before and after the experiment? Did it vary by potato type?
2. What does the net change in the potato sample indicate?
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Osmosis
3. Different types of potatoes have varying natural sugar concentrations. Explain how this may influence the water potential of each type of potato.
4. Based on the data from this experiment, hypothesize which potato has the highest natural sugar concentration. Explain your reasoning.
5. Did water flow in or out of the plant cells (potato cells) in each of the samples examined? How do you know this?
6. Would this experiment work with other plant cells? What about with animal cells? Why or why not?
7. From what you know of tonicity, what can you say about the plant cells and the solutions in the test tubes? 8. What do your results show about the concentration of the cytoplasm in the potato cells at the start of the experiment? 9. If the potato is allowed to dehydrate by sitting in open air, would the potato cells be more likely to absorb more or less water? Explain.
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Lab 8
Enzymes
Enzymes
Learning Objectives
•
Describe enzyme structure and function including activation site
•
Explain how enzymes affect activation energy of a reaction
•
Relate reaction kinetics, activators and inhibitors to the function of enzymes
•
Hypothesize how climate-based changes can affect enzyme function
Introduction
Enzymes are specialized proteins made by living cells that serve as biological catalysts. A catalyst serves to decrease the activation energy normally needed for a reaction to occur naturally. Enzymes can increase the reaction rate between two molecules by over a million times faster than it would be without the enzyme.
Most biochemical reactions require enzymes for them to occur at fast enough rates to be useful. Typical nomenclature for enzymes uses name of the substrate or the chemical reaction it catalyzes as the prefix, and adds the segment “-ase” as the suffix (e.g., catalase, amylase, polymerase, etc.). Keep in mind that some enzymes, such as pepsin, do not follow this nomenclature.
Enzymes are extremely selective, and are often described as adhering to a “lock and key fit” (Figure 1). Their selectivity is primarily enforced by their molecular shape, hydrophilic interactions, and electrostatic forces.
These factors dictate which substrate(s) the enzyme can bind and affect. The activation site is a region on the enzyme where the substrate attaches to form the enzyme/substrate complex. This is also where the reaction occurs. After the enzyme/substrate complex forms and catalysis occurs, the modified substrate is released from the active site, and the enzyme can repeat the process. Enzymes levels are not reduced or altered during the reaction. This means they are efficient and can be used repeatedly.
Figure 1: Demonstration of the “lock and key” fit. The empty pie slice located at the top of Part
A reflects the active site. Part B displays the substrate-enzyme complex. Part C displays the release of the modified substrate, or products.
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Enzymes
Activation Energy
Molecules harbor energy within their chemical bonds. This energy is released when the bonds are broken, thus defining one of the primary reasons why bonds are broken in biological pathways such as metabolism.
However, chemical reactions require an initial amount of energy to break and form bonds (even if the reaction is exothermic). This is often referred to as activation energy as it corresponds to the minimum amount of energy needed to engage (or, activate) a reaction. Enzymes are useful in chemical reactions as they can lower the activation energy required to initiate the reaction. For example, consider trying to roll a large rock on a flat surface. This can be very difficult to do; however, what if you only had to roll the rock down a hill? This would make the process much easier to accomplish. Enzymes are homologous to the downhill slope by decreasing the effort it takes to start a chemical reaction.
Activation energy makes molecules unstable. If sufficient activation energy is available, the bonds existing within the now unstable molecule break to create new, more stable molecules. Activation energy separates the energy levels of the substrates and products and by amounts which are sufficient to initiate the transition state (in which the reactants become unstable) so the reaction can occur. Energy must be added to the substrates to overcome this barrier, which can be recovered when products are formed.
Reaction Kinetics
Enzymes influence the rate at which a chemical reaction occurs. Their activity is affected by three primary factors:
•
Salt concentration
•
Temperature
•
pH
Salt concentrations in the enzyme’s environment, either too high or too low, can influence the behavior of the charged amino acids that make up the protein structure of the enzyme. When this occurs, normal interaction of the charged groups is blocked and the enzyme is no longer functional.
Temperature increases the rate of chemical reactions in two different ways: increasing or decreasing the reaction rate constant and thermal denaturation.
The reaction rate constant can be increased or decreased, and typically adheres to a linear relationship to the direction in which the temperature is adjusted. However, if the temperature rises above an optimum value, enzymes or (their substrates) may become damaged, lose functionality or denature entirely. The pH can influence the efficiency or potential of an enzyme to perform. In fact, the majority of enzymes can only function within a narrow pH range of
114
?
Did You Know...
Enzymes, including proteases and amylases, started being used in laundry detergents in
1913. At the time, this integration was not economically practical, and the enzymes stained the machines used to process the detergent. However, almost 50% of liquid detergents and 25% of powdered detergents now incorporate enzymes to help break down stains which can’t be removed with chemical surfactants alone.
Enzymes approximately 5 - 9. Specifically, pH affects reaction kinematics through the following techniques:
1. Altering the binding site on the enzyme.
2. Changing the protein structure of the enzyme itself
3. Revising the ionization status of the enzyme or substrate.
How do you think the functional pH of enzymes in the stomach compares to the optimal pH for enzymes in blood?
The concentrations of both the enzyme and substrate can also influence the reaction rate (Figure 2). The maximum velocity of the reaction is achieved at a saturating concentration of substrate, at which every enzyme molecule is utilized in the reaction.
?
Did You Know...
At low substrate concentrations, an enzyme can readily convert a substrate into products. However, as more substrate is added, an enzyme will reach a saturation point at which the reaction rate will plateau.
This point occurs when there are more substrate molecules than the enzyme can convert in a given period of time. This interaction falls into the category of
“saturation kinetics”.
Figure 2: Reaction rate increases with higher substrate concentrations until it meets maximum velocity.
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Enzymes
Cofactors: Activators and Inhibitors
Substances other than the substrate, called enzyme co-factors, can bind to the enzyme and influence its behavior. Unlike enzymes, coenzymes bind to the enzyme and are structurally modified during the chemical reaction. There are two primary category of cofactors: activators and inhibitors. Activators are chemicals that bind to the active site of the enzyme and help it to bind to the substrate. They are sometimes called cofactors or organic coenzymes.
Inhibitors are chemicals that interfere with the binding of the substrate to the enzyme. There are two types;
1. Competitive: Cannot be replaced by the substrate.
2. Non-competitive: Can not removed by the substrate.
?
Did You Know...
Many drugs and poisons are enzyme inhibitors. For example, aspirin inhibits an enzyme that leads to inflammation. Normal cellular processes produce toxic substances (waste) such as hydrogen peroxide and free radicals that will kill the cell if they are not eliminated. Fortunately, yeast and other organisms (including humans) have an enzyme called catalase that breaks down hydrogen peroxide into oxygen and water, which are both harmless to cells. Thus, enzymes are responsible for both constructive
(anabolism) and destructive (catabolism) activities that work synergistically to pattern the overall metabolism of an organism.
Pre-Lab Questions
1. How could you test to see if an enzyme was completely saturated during an experiment?
2. List three conditions that would alter the activity of an enzyme. Be specific with your explanation.
3. Take a look around your house and identify household products that work by means of an enzyme. Name the products, and indicate how you know they work with an enzyme.
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Enzymes
Experiment 1: Enzymes in Food
Amylase is used by humans to facilitate digestion. Specifically, it is an enzyme which breaks down starch molecules into sugar molecules. This is why people sometimes observe a sweet taste after sucking on a starch-containing food for an extended period of time. Amylase is found naturally in human saliva and the pancreas. However, it is also present in some of the common plant foods consumed by humans.
This experiment tests for the presence of amylase in food by using Iodine-Potassium Iodide, IKI. IKI is a color indicator used to detect starch. This indicator turns dark purple or black in color when in the presence of starch. Therefore, if the IKI solution turns to a dark purple or black color during the experiment, one can determine that amylase is not present (because presence of amylase would break down the starch molecules, and the IKI would not change color).
Materials
(1) 2 oz. Bottle (Empty)
(1) 100 mL Graduated Cylinder
*2 Food Products (e.g., ginger root, apple, potato, etc.) 30 mL Iodine-Potassium Iodide, IKI
*Kitchen Knife
Permanent Marker
*Paper Towel
Ruler
*Saliva Sample
2 Spray Lids
*Tap Water
30 mL Liquid Starch, C6H10O5
*Cutting Board
*You Must Provide
Procedure:
1. Remove the cap from the starch solution. Attach the spray lid to the starch solution.
2. Rinse out the empty two ounce bottle with tap water. Use the 100 mL graduated cylinder to measure and pour 30 mL of IKI into the empty two ounce bottle. Attach the remaining spray lid to the bottle.
3. Set up a positive control for this experiment by spraying a paper towel with the starch solution. Allow the starch to dry for approximately one hour (this time interval may vary by local humidity).
4. In the mean time, set up a negative control for this experiment. Use your knowledge of the scientific method and experimental controls to establish this component (hint: what should happen when IKI solution contacts something that does not contain starch?) Identify your negative control in Table 1.
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Enzymes
Note: Be sure to space the positive and negative controls apart from each other to prevent cross contamination.
5. When the starch solution has dried, test your positive and negative controls by spraying them with IKI solution. This step establishes a baseline color scale for you to evaluate the starch concentration of the food products you will test in Steps 7 - 11. Record your results in Table 1.
6. Select two food items from your kitchen cabinet or refrigerator.
7. Obtain a kitchen knife and a cutting board. Carefully cut your selected food items to create a fresh surface.
8. Gently rub the fresh/exposed area of the food items on the dry, starch-sprayed paper towel back and forth 10 - 15 times. Label where each specimen was rubbed on the paper towel with a permanent marker (Figure 3).
9. Wash your hands with soap and water.
10. Take your finger and place it on your tongue to transfer some saliva to your finger. Then, rub your moistened finger saliva into the paper towel. Repeat this step until you are able to adequately moisten the paper towel.
Figure 3: Sample set-up.
Note: You should always wash your hands before touching your tongue! Alternatively, if you do not wish to put your hands in your mouth, you may also provide a saliva sample by spitting in a separate bowl and rubbing the paper towel in the saliva. Be sure not to spit on the paper towel directly as you may unintentionally cross-contaminate your samples.
11. Wait five minutes.
12. Hold the IKI spray bottle 25 - 30 cm away from the paper towel, and mist with the IKI solution.
13. The reaction will be complete after approximately 60 seconds. Observe where color develops, and consider what these results indicate. Record your results in Table 1.
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Enzymes
Table 1: Substance vs. Starch Presence
Substance
Resulting Color
Presence of Starch?
Positive Control: Starch
Negative Control: Student Must Select
Food Product:
Food Product:
Saliva:
Post-Lab Questions
1. What were your controls for this experiment? What did they demonstrate? Why was saliva included in this experiment?
2. What is the function of amylase? What does amylase do to starch?
3. Which of the foods that you tested contained amylase? Which did not? What experimental evidence supports your claim?
4. Saliva does not contain amylase until babies are two months old. How could this affect an infant’s digestive requirements?
5. There is another digestive enzyme (other than salivary amylase) that is secreted by the salivary glands.
Research to determine what this enzyme is called. What substrate does it act on? Where in the body does it become activated, and why?
6. Digestive enzymes in the gut include proteases, which digest proteins. Why don’t these enzymes digest the stomach and small intestine, which are partially composed of protein?
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Enzymes
Experiment 2: Effect of Temperature on Enzyme Activity
Yeast cells contain catalase, an enzyme which helps convert hydrogen peroxide to water and oxygen. This enzyme is very significant as hydrogen peroxide can be toxic to cells if allowed to accumulate. The effect of catalase can be seen when yeast is combined with hydrogen peroxide (Catalase: 2
2 H2O + O2).
H2O2
In this lab you will examine the effects of temperature on enzyme (catalase) activity based on the amount of oxygen produced. Note, be sure to remain observant for effervescence when analyzing your results.
Figure 4: Catalase catalyzes the decomposition of hydrogen peroxide to water and oxygen.
Materials
3 Balloons
3 Test Tubes (Glass)
(2) 250 mL Beakers
Test Tube Rack
30 mL 3% Hydrogen Peroxide, H2O2
Thermometer
Measuring Spoon
Yeast Packet
Permanent Marker
Ruler
*Hot Water Bath
*Stopwatch
20 cm String
*You Must Provide
Procedure
1. Use a permanent marker to label test tubes 1, 2, and 3. Place them in the test tube rack.
2. Fill each tube with 10 mL hydrogen peroxide. Then, keep one of the test tubes in the test tube rack, but transfer the two additional test tubes to two separate 250 mL beakers.
3. Find one of the balloons, and the piece of string. Wrap the string around the uninflated balloon and measure the length of the string with the ruler. Record the measurement in Table 2.
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Enzymes
4. Create a hot water bath by performing the following steps:
a. Determine if you will use a stovetop or microwave to heat the water. Use the 100 mL graduated cylinder to measure and pour approximately 200 mL of water into a small pot or microwave safe bowl (you will have to measure this volume in two separate allocations).
b. If using a stovetop, obtain a small pot and proceed to Step 4c. If using a microwave, obtain a microwave-safe bowl and proceed to Step 4e.
c. If using a stove, place a small pot on the stove and turn the stove on to a medium heat setting.
d. Carefully monitor the water in the pot until it comes to a soft boil (approximately 100 °C). Use the thermometer provided in your lab kit to verify the water temperature. Turn the stove off when the water begins to boil. Immediately proceed to Step 5.
CAUTION: Be sure to turn the stove off after creating the hot water bath. Monitor the heating water at all times, and never handle a hot pan without appropriate pot holders.
e. If using a microwave, place the microwave-safe bowl in the microwave and heat the water in 30 second increments until the temperature of the water is approximately 100 °C. Use the thermometer provided in your lab kit to verify the water temperature. Wait approximately one minute before proceeding to Step 5.
5. Place Tube 1 in the refrigerator. Leave Tube 2 at room temperature, and place Tube 3 in the hot water bath. Important Note: The water should be at approximately 85 °C when you place Tube 3 in it. Verify the temperature with the thermometer to ensure the water is not too hot! Temperatures which exceed approximately 85 °C may denature the hydrogen peroxide.
6. Record the temperatures of each condition in Table 2. Be sure to provide the thermometer with sufficient time in between each environment to avoid obscuring the temperature readings.
7. Let the tubes sit for 15 minutes.
8. During the 15 minutes prepare the balloons with yeast by adding 1/4 tsp. of yeast each balloon. Make sure all the yeast gets settled to the bulb of the balloon and not caught in the neck. Be sure not spill yeast while handling the balloons.
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Enzymes
9. Carefully stretch the neck of the balloon to help ensure it does not rip when stretched over the opening of the test tube.
10. Attach the neck of a balloon you prepared in Step 8 to the top of Tube 2 (the room temperature test tube) making sure to not let the yeast spill into the test tube yet. Once the balloon is securely attached to the test tube lift the balloon and allow the yeast to enter the test tube. Tap the bulb of the balloon to ensure all the yeast falls into the tube.
11. As quickly and carefully as possible remove the Tube 1 (cold) from the refrigerator and repeat steps 9 10 with Tube 1 using a balloon you prepared in Step 8.
12. As quickly and carefully as possible remove Tube 3 (hot) from the hot water bath and repeat steps 9 - 10 with Tube 3 using a balloon you prepared in Step 8.
13. Swirl each tube to mix, and wait 30 seconds.
14. Wrap the string around the center of each balloon to measure the circumference. Measure the length of string with a ruler. Record your measurements in Table 2.
Table 2: Balloon Circumference vs. Temperature
Tube
Temperature (°C)
Balloon Circumference
(Uninflated; cm)
Balloon Circumference
(Final; cm)
1 - (Cold)
2 - (RT)
3 - (Hot)
Post-Lab Questions
1. What reaction is being catalyzed in this experiment?
2. What is the enzyme in this experiment? What is the substrate?
3. What is the independent variable in this experiment? What is the dependent variable?
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Enzymes
4. How does the temperature affect enzyme function? Use evidence from your data to support your answer.
5. Draw a graph of balloon diameter vs. temperature. What is the correlation?
6. Is there a negative control in this experiment? If yes, identify the control. If no, suggest how you could revise the experiment to include a negative control.
7. In general, how would an increase in substrate alter enzyme activity? Draw a graph to illustrate this relationship.
8. Design an experiment to determine the optimal temperature for enzyme function, complete with controls.
Where would you find the enzymes for this experiment? What substrate would you use?
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Lab 9
Cellular Respiration
Cellular Respiration
Learning Objectives
•
Compare and contrast aerobic respiration to anaerobic respiration including the role of ADP and
ATP
•
Relate glucose to pyruvate during the process of glycolysis
•
Explain the role of the citric acid cycle and oxidative phosphorylation in the process of aerobic respiration
•
Explain the role of fermentation in the process of anaerobic respiration
Introduction
Cellular respiration refers to the process of converting chemical energy existing within organic molecules, such as sugars, into a form which can be immediately used by organisms. This process is required for both aerobic (organisms which have oxygen available) and anaerobic (organisms which do not have oxygen available) organisms, but it is executed differently based on the compounds and structures local to the host. However, both processes share the common goal of harvesting biological energy stored in fuel molecules (such as glucose) and convert this energy into ATP. The reaction is written as follows:
C6H12O6
(glucose)
+
6 O2
6 CO2
+
6 H2O +
(oxygen)
(carbon dioxide)
energy
(water)
ADP and ATP
Adenosine triphosphate, or ATP, is often referred to as the energy currency of the cell. It is used in the majority of cellular functions including active transport across a membrane, cell structure maintenance, DNA and RNA synthesis, protein synthesis, etc. However, even though this molecule is extremely important, sometimes more ATP is required than is available.
When ATP levels become too low, a special protein signals the cell to begin respiration.
Respiration occurs in anaerobic and aerobic organisms, therefore, there is more than one pathway used to produce ATP.
However, a high level analysis can conclude that in both types of organisms, insufficient concentrations of ATP can trigger a series of metabolic reactions which ultimately replenish the local
ATP supply. These reactions are primarily catabolic; in other
Figure 1: ATP (above) consists of the nucleotide adenine, a 5-sided sugar (ribose), and three phosphates. Each is bound to three oxygen molecules. Cleavage of one phosphate molecule from ATP yields the amount of energy (calories) found in one peanut.
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Cellular Respiration words, they break down big molecules into smaller pieces. Free energy is released when the molecules are broken down, which can be sequentially used in each proceeding step. Through a series of oxidation reactions, the cell converts carbohydrates into carbon dioxide and water. It also transforms adenosine diphosphate (ADP) into ATP by adding a free phosphate molecule to ADP. This set of reactions provides a constant source of energy for the cell, as long as all the critical components for the reactions are available.
Glycolysis
The first stage of respiration is called glycolysis. Glycolysis occurs with or without oxygen and takes place in the cytoplasm. During glycolysis, glucose is broken down to provide energy for the cell. The glycolytic pathway is highly conserved amongst organisms, and is found in all living organisms.
Glucose contains high energy bonds which release electrons (energy) when broken. Through this process, the six-carbon glucose molecule is broken down into two, three-carbon molecules called pyruvic acid
(pyruvate). Though the bonds holding pyruvate together contain a great deal of potential energy, this step yields little energy. Only two ATP molecules and two nictotinamide adenine dinucleotide (NADH) molecules are produced during glycolysis.
Aerobic Respiration vs. Fermentation
The next stage of respiration depends on if the organism is aerobic or anaerobic; or, in the case of multicellular organisms such as plants, if oxygen is available to the cell. In an aerobic environment, the citric acid cycle begins. Conversely, if an anaerobic environment exists, fermentation or anaerobic respiration takes place.
Plants primarily engage in aerobic respiration. However, in rare cases such as extremely wet soil which results in waterlogged roots, anaerobic respiration may also occur in plants. The steps involved in aerobic respiration, anaerobic respiration, and fermentation vary, but all involve the manipulation of electrons and serve to perpetuate the availability of energy for the cell.
In aerobic respiration, pyruvate molecules are shuttled from the cytoplasm into the mitochondria to prepare for the citric acid cycle. An enzyme complex called pyruvate dehydrogenase complex produces acetylcoenzyme A (acetyl-CoA) by initially breaking down pyruvate to a two-carbon acetyl group and carbon dioxide. The acetyl group is then transferred to coenzyme A. Coenzyme A then acts as a carrier for the twocarbon acetyl group and transfers the acetyl group to the citric acid cycle.
The Citric Acid Cycle
The citric acid cycle, also known as the Krebs cycle or tricarboxylic acid cycle (TCA cycle), is an eight step process in which the acetyl group is oxidized to carbon dioxide. The citric acid cycle engages both anabolic and catabolic reactions to construct and break down large molecules. During this process, two important electron shuttle molecules [flavin adenine dinucleotide (FAD) and nicotinamide adenine dinucleotide (NAD+)] are reduced forming FADH2 and NADH. These molecules shuttle electrons to the next step of aerobic respiration, the electron transport chain.
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Cellular Respiration
The electron transport chain, ETC, performs the final series of biochemical reactions. Ultimately, the
ETC functions to regenerate oxidized molecules (coenzymes) from their reduced state so that other glucose molecules can be converted to energy through future rounds of respiration. To accomplish this objective, electrons (and hydrogen ions) from FADH2 and NADH are transferred to enzyme complexes embedded in the mitochondria’s inner membrane. These complexes act as electron acceptors, passing the electrons from one complex to the next. Since oxygen has a very high affinity for electrons, aerobic respiration is the most efficient means of producing ATP (up to 34 molecules are generated per round).
Figure 2: The electron transport chain.
Fermentation
Eukaryotic and prokaryotic organisms use fermentation in anaerobic conditions. In these cases, the citric acid cycle and the ETC are not used. Instead, pyruvate is the final substrate. These molecules are metabolized in the cytoplasm to produce alcohol or lactic acid, depending on the organism. The main function of fermentation is the reduction of NADH to NAD+. This contributes to the propagation of the glycolytic pathway, which allows ATP production to continue. Recall that glycolysis nets two ATP molecules per glucose molecule. Fermentation allows for the generation of much less ATP per glucose molecule than aerobic respiration because the citric acid cycle and the ETC are not engaged.
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Did You Know...
Yeast has been used to make leavened bread for centuries. When yeast undergoes fermentation, carbon dioxide is trapped between gluten and causes the bread to rise. Ethanol, another byproduct of yeast fermentation, generates the alcohol content in beer, and the carbon dioxide provides effervescence.
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Cellular Respiration
Figure 3: Anaerobic vs. aerobic respiration charts. The green area represents the cytoplasm of the cell where glycolysis and anaerobic respiration occur. The blue area represents the inside of the mitochondria where aerobic respiration occurs.
Industrial Applications
Fermentation has many industrial applications, many of which are evident within the food industry. For example, the sour or tart taste of yogurt and sauerkraut is due to bacteria in the Lactobacillus genus. Lactobacillus perform lactate fermentation in which pyruvic acid (from glycolysis) is reduced directly to the final end product, lactic acid. Another example of industrial fermentation focuses on yeast. Yeast, a unicellular eukaryote, has been used to make leavened (rising) bread for centuries. When yeast undergoes fermentation, carbon dioxide is trapped between gluten and causes the bread to rise. Ethanol is another byproduct of fermentation and is responsible for the alcohol content in beer. Carbon dioxide produced through fermentation is responsible for the effervescence in beer.
Respirometers
Scientists may use respirometers to measure the rate of respiration. This device capitalizes on the ideal gas law for its function. The ideal gas law applies to those gases in which all collisions between atoms or molecules are perfectly elastic and no intermolecular forces exist. The internal energy is completely kinetic, and any change in energy will be accompanied by a change in temperature, volume, and pressure. The relationship is defined by the ideal gas law equation:
PV = nRT
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Cellular Respiration
In this equation, P is the absolute pressure, V is the volume, R is the universal gas constant (8.3145 J/mol °
K), and T is the absolute temperature (Kelvin). Respirometers typically consist of a device to absorb carbon dioxide produced through respiration and measure oxygen uptake by displacement of fluid in a tube connected to the sealed chamber. Since the carbon dioxide is absorbed, air will be taken in through the tube
(displacing the fluid) to maintain equal pressure within the chamber.
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Did You Know...
Cells require more energy during physical activity than when the body is at rest. Humans rely on aerobic respiration to provide this energy. However, some physical activities (such as a strenuous workout) may create an anaerobic environment if the oxygen supply depletes before the demand is removed. When this occurs, lactate fermentation will begin to provide additional energy without the use of oxygen. Lactic acid is a 3-carbon molecule. Therefore, most of the energy remains and the only energy produced is the two ATP molecules generated from glycolysis. The buildup of lactic acid can cause muscles to feel sore and stiff. This feeling persists until the lactic acid is removed from the muscles by diffusion into the blood.
Definitions
• Aerobic Respiration: Oxidation of molecules to produce energy in the presence of oxygen.
•
Anaerobic Respiration: Oxidation of molecules to produce energy in the absence of oxygen.
•
ATP: Adenosine triphosphate; the main energy source of the cell.
•
Citric Acid Cycle (The Krebs Cycle): The step following glycolysis in which acetyl groups produced using pyruvate molecules are oxidized yielding CO2, ATP, H2O and reduced electron shuttle molecules.
•
Fermentation: A metabolic pathway used in anaerobic respiration which reduces NADH to NAD+
•
Glycolysis: The initial process of energy metabolism for anaerobic and aerobic organisms which produces two ATP, two NADH, and two pyruvate molecules.
•
Oxidation: The loss of electrons.
•
Oxidation State: The degree of oxidation of an atom.
•
Pyruvate (Pyruvic Acid): A three-carbon molecule generated through glycolysis.
•
Reduction: Gain of electrons.
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Cellular Respiration
Pre-Lab Questions
1. Why is cellular respiration necessary for living organisms?
2. Why is fermentation less effective than respiration?
3. What is the purpose of glycolysis?
4. How many ATP molecules are produced in aerobic respiration, fermentation, and glycolysis?
Experiment 1: Fermentation by Yeast
Yeast cells produce ethanol, C2H6O, and carbon dioxide, CO2, during alcoholic fermentation. In this experiment, you will measure the production of CO2 to determine the rate of anaerobic respiration in the presence of different carbohydrates with a simplified respirometer.
Materials
(4) 250 mL Beakers
Ruler
15 mL 1% Glucose Solution
15 mL 1% Sucrose (Sugar) Solution, C12H22O11
(1) 100 mL Graduated Cylinder
Test Tube Rack
Measuring Spoon
1 Yeast Packet
1 g. Packets of Equal®, Splenda®, and Sugar
*Stopwatch
Permanent Marker
*Warm Water
3 Pipettes
5 Respirometers (two test tubes that fit into each other – 5 plastic and 5 glass; see Figure 4).
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* You Must Provide
Cellular Respiration
Note: Sucrose (a disaccharide) is made up of glucose and fructose. Glucose is a monosaccharide.
Figure 4: Four respirometers. Note how the smaller, plastic test tube is inverted into the larger test tube. You will create five respirometers in this experiment.
Procedure
1. In this experiment, you will mix yeast with sugar, Equal, and Splenda. Before you begin, develop a hypothesis predicting what will happen when the sugar/sweeteners are mixed with yeast. Example, will fermentation occur? Why or why not? Record your hypothesis in the Post-Lab Questions section.
2. Use the permanent marker to label three 250 mL beakers as Equal®, Splenda®, and Sugar.
3. Empty the Equal®, Splenda®, and Sugar packets into the corresponding beakers.
4. Fill the Equal® and Splenda® beakers to the 100 mL mark with tap water.
5. Fill the Sugar beaker to the 200 mL mark.
6. Mix each beaker thoroughly by pipetting the solution up and down. Each beaker now contains a 1% solution. Set these aside for later use.
7. Completely fill the smallest tube with tap water and invert the larger tube over it. Push the small tube up
(into the larger tube) until the top connects with the bottom of the inverted tube. Invert the respirometer so that the larger tube is upright (there should be a small bubble at the top of the internal tube).
Note: Repeat Step 6 several times as practice. Strive for the smallest bubble possible. When you feel comfortable with this technique, empty the test tube(s) and proceed to Step 7.
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Cellular Respiration
8. Use the permanent marker to label the fourth 250 mL beaker as Yeast.
9. Fill this beaker with 175 mL of warm tap water. The exact temperature does not matter, but the water should be warm to the touch.
10. Open the yeast package, and use the measuring spoon to measure and pour 1 tsp. yeast into the beaker. Pipette the solution up and down until all of the yeast has dissolved into solution.
Note: Make sure the yeast solution remains homogenous before each test tube is filled in the proceeding steps. The yeast concentration is fairly saturated, and yeast may precipitate out of solution if the beaker rests for an extended period of time.
11. Use the permanent marker to label the big and small test tubes as 1, 2, 3, 4, and 5.
12. Use the 100 mL graduated cylinder to measure and pour 15 mL of the following solutions into the corresponding small test tubes:
Tube 1: 1% Glucose Solution
Tube 2: 1% Sucrose Solution
Tube 3: 1% Equal® Solution
Tube 4: 1% Splenda® Solution
Tube 5: 1% Sugar Solution
Note: Rinse the graduated cylinder between each measurement.
13. Fill the remaining volume in each small tube to the top with the yeast solution.
14. Slide the corresponding larger tube over the small tube and invert it as practiced in Step 6. This will mix the yeast and sugar/sweetener solutions.
15. Place the respirometers in the test tube rack, and use a ruler measure the initial air space in the rounded bottom of the internal tube. Record these values in the Table 1.
16. Allow the test tubes to sit in a warm place (approximately 30 °C) for two hours. Placement suggestions include: a sunny windowsill, atop (not in!) a warm oven heated to approximately 85 °C (185 °F on an oven setting), or under a very bright (warm) light.
17. At the end of the respiration period, use your ruler to measure the final gas height (total air space) in the tube. Record this data in Table 1.
Table 1: Yeast Fermentation Data
Tube
134
Initial Gas Height (mm)
Final Gas Height (mm)
Net Change
Cellular Respiration
Post-Lab Questions
1. Include your hypothesis from Step 1 here. Be sure to include at least one piece of scientific reasoning in your hypothesis to support your predictions.
2. Did you notice a difference in the rate of respiration between the various sugars? Did the artificial sugar provide a good starting material for fermentation?
3. Was anaerobic fermentation occurring? How do you know (use scientific reasoning)?
4. If you observed respiration, identify the gas that was produced. Suggest two methods you could use for positively identifying this gas.
5. Hypothesize why some of the sugar or sweetener solutions were not metabolized, while others were. Research the chemical formula of Equal® and Splenda® and explain how it would affect yeast respiration.
6. How do the results of this experiment relate to the role yeast plays in baking?
7. What would you expect to see if the yeast cell metabolism slowed down? How could this be done?
8. Indicate sources of error and suggest improvement (for example, what types of controls could be added?).
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Cellular Respiration
Experiment 2: Aerobic Respiration in Beans
We will evaluate respiration in beans by comparing carbon dioxide production between germinated and nongerminated beans. As shown in the balanced equation for cellular respiration, one of the byproducts is CO2
(carbon dioxide):
C6H12O6 + 6 H2O + 6 O2
energy + 6 CO2 +12 H2O
We will use a carbon dioxide indicator ( bromothymol blue) to show oxygen is being consumed and carbon dioxide is being released by the beans. Bromothymol blue is an indicator that turns yellow in acidic conditions, green in neutral conditions, and blue in basic conditions. When carbon dioxide dissolves in water, carbonic acid is formed by the reaction:
H2O + CO2
H2CO3
resulting in the formation of this weak acid. If an indicator such as bromothymol blue is present, what do you think would happen? (Hint - what color would the indicator change to?)
Materials
(6) 250 mL Beakers
1 Pipette
24 mL Bromothymol Blue Solution, C27H28Br2O5S
6 Rubber Bands (Large. Contains latex; please handle with gloves if you have a latex allergy.)
100 Kidney Beans
6 Medicine Cups (small, clear, plastic cups)
15 cm Parafilm®
*Water
*Paper Towels
Permanent Marker
100 Pinto Beans
*You Must Provide
Procedure
1. Label two of the 250 mL beakers as Soaked: Pinto and Soaked: Kidney.
2. Fill each beaker with 200 mL water.
3. Count and transfer 50 pinto beans into the Soaked: Pinto beaker. Then, count and transfer 50 kidney beans into the Soaked: Kidney beaker. Allow the beakers to rest for 24 hours.
4. After 24 hours have passed, carefully strain the water from each beaker.
5. Place two paper towels on a flat work surface. Use the permanent marker to label one paper towel as
Soaked: Pinto and the second as Soaked: Kidney.
6. Pour the soaked beans onto paper towels, keeping them sorted by bean type.
7. Label the remaining beakers as Dry: Pinto, No Pinto, Dry: Kidney, and No Kidney.
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Cellular Respiration
8. Place several layers of moist paper towels at the bottom of all six, 250 mL beakers.
9. Place 50 pre-soaked pinto beans into the Soaked: Pinto beaker, 50 unsoaked pinto beans into the Dry:
Pinto beaker, and no beans into the No Pinto beaker.
10. Place 50 pre-soaked kidney beans into the Soaked: Kidney, 50 unsoaked kidney beans into the Dry: Kidney beaker, and no beans into the No Kidney beaker.
11. Dispense four mL of bromothymol blue solution into each of the six measuring cups. Then, place one cup inside each beaker (Figure 5).
12. Stretch Parafilm® across the top of each beaker. Secure with a rubber band to create an air-tight seal.
Note: If your Parafilm® seal breaks, plastic wrap
(such as Saran® wrap) can be used as a replacement.
13. Place the beakers on a shelf or table, and let sit undisturbed at room temperature.
14. Record the initial color of the bromothymol solutions and observe the jars at 30 minute intervals for three hours. Record the color of the bromothymol blue in
Tables 2 and 3.
Figure 5: The image above shows what the beans,
15. Let the beans and the jar sit overnight. Record your beaker, and measuring cup set-up should look like. final observations in Tables 2 and 3.
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Cellular Respiration
Table 2: Bromothymol Blue Color Change Over Time for Pinto Bean Trial
Time
Pre-Soaked Pinto Beans
Dry Pinto Beans
No Pinto Beans
0 min
30 min
60 min
90 min
120 min
150 min
180 min
24 hours
Table 3: Bromothymol Blue Color Change Over Time for Kidney Bean Trial
Time
0 min
30 min
60 min
90 min
120 min
150 min
180 min
24 hours
138
Pre-Soaked Kidney Beans
Dry Kidney Beans
No Kidney Beans
Cellular Respiration
Post-Lab Questions:
1. How did the color of the bromothymol blue solution in each beaker change over time in each condition?
2. What is the mechanism driving the bromothymol blue solution color change?
3. What can be inferred from the color change of the bromothymol blue solution?
4. What evidence do you have to prove that cellular respiration occurred in the beans? Explain your answer.
5. What are the controls in this experiment, and what variables do they eliminate? Why is it important to have a control for this experiment?
6. If this experiment were conducted at 0 °C, what difference would you see in the rate of respiration? Why?
7. Would you expect to find carbon dioxide in your breath? Why?
8. What else could you incorporate into this experiment to verify that the gas is responsible for the color change? Design an experiment that shows the steps required.
139
Lab 10
Cell Structure and Function
Cell Structure and Function
Learning Objectives
•
Apply Cell Theory
•
Compare and contrast the structure and function of prokaryotic and eukaryotic cells
•
Identify eukaryotes and prokaryotes based on their cellular structure
Introduction
A cell is the fundamental unit of life. All living organisms originate from a single cell. Some remain as a single cell, while others grow and divide to form a multi-cellular organism. Though most cells are difficult to see with the naked eye, cytologists have successfully identified many of their features using microscopes. The scope of these features ranges from the fundamental characteristics of the outer membranes, to the internal structures nestled within the cytoplasm. This structural information provided the foundation for much of modern biology, and formed the basis of Cell Theory.
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Cell Theory states:
• All living things are made of cell(s).
• A cell is the smallest unit of life.
Did You Know...
Cytologists are scientists who study cells.
The study of the cell is known as cytology.
• All cells are generated from previous cells.
• All cells pass on genetic information.
• Energy metabolism occurs within cells.
• The chemical make-up of all cells is similar (but not identical).
Functional Requirements of Life
The functional requirements of life are similar to the content presented in Cell Theory. In essence, these requirements state that all living organisms (unicellular or multicellular) must perform the following functions:
1. Movement: Cells can change position or shape.
2. Responsiveness: Cells can sense internal or external changes; and, react to these changes.
3. Respiration/Metabolism: Cells produce energy through cellular chemical reactions
4. Reproduction: Cells are able to produce offspring.
5. Excretion: Cells can remove waste produced by metabolic activities.
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Cell Structure and Function
Types of Cells: Prokaryotes and Eukaryotes
Although all organisms are made up of one or more cells, not all cells are identical. Prokaryotes and eukaryotes are organisms composed of two structurally different types of cells. This division offers the most broad distinction between different types of organisms. Prokaryotes are the most ancient and basic organisms, and span the taxonomic classes of bacteria and archaea. They lack a membrane-bound nucleus and membranebound organelles (specialized structures). The term prokaryote comes from the Latin words “pro” (before) and
“karyote” (nucleus).
Eukaryotes are much more complex organisms with two characteristics that set them apart from prokaryotes: a defined nucleus, and membrane-bound organelles. The term “eukaryote” comes from the Latin words
“eu” (true) and “karyote” (nucleus). Fungi, plant, protist, and animal cells are all examples of eukaryotic cells.
Prokaryotic and Eukaryotic Structural Similarities
While the overall structure of prokaryotic and eukaryotic cells differs, there are several structures which are commonly found in all cells. For example, all cells have deoxyribonucleic acid (DNA), cytoplasm, a cell membrane, and ribosomes.
DNA
DNA contains all the genetic information for the cell. It is made up of two long chains of nucleotides. Each nucleotide consists of a base, a five carbon sugar molecule, and at least one phosphate group. Adenine, thymine, guanine, and cytosine are the four different types of nucleotides. These nucleotides connect to form a sugar phosphate backbone, and extend hydrogen bonds out towards complimentary DNA strands to form a double helix structure. The double helix is often compared to a long, twisted ladder. DNA is often found as a DNA-protein complex known as chromatin. Chromatin condenses during cell cycle division to form chromosomes.
G
C
A
T
C
G
T
A
Figure 1: The hydrogen bonds formed
All cells have the DNA described, although eukaryotic DNA is linear, between adenine and thymine and while prokaryotic DNA is circular. Also, because prokaryotes do not cytosine and guanine contribute to have a nucleus, their DNA exists freely within the nucleoid region as the double-helix structure of DNA opposed to the eukaryotic location within the nucleus.
Cytoplasm
Cytoplasm is a fluid-like substance which also exists in all cells. The cytoplasm is made up cytosol, a water based solution containing small molecules, such as proteins and sugars. Because of the abundance of small
144
Cell Structure and Function molecules and particles, the cytoplasm is better understood as a thicker, gel-like solution instead of a highly viscous liquid solution. All of the everyday cellular functions are carried out in the cytoplasm, including cell replication and cell growth.
There are also many biomolecules, such as ribosomes, that are suspended within the cytoplasm. Ribosomes are particles made up of RNA and proteins and are roughly 20 nm (typical prokaryotic length) to 30 nm (typical eukaryotic length) in diameter. These particles are the site for the complex process of protein synthesis in all cells.
Cell Membrane
The cytoplasm is surrounded by a cell membrane. The cell membrane provides structural support and strength. It also protects the cell and the cellular content from the external environment. It acts as a selective gateway which regulates the flow of atoms and molecules in and out of the cell. The cell membrane can be described by the fluid mosaic model which states that cell membranes are not static. Instead, they express movement by the molecules that compose it. Cell membranes are often referred to as a phospholipid bilayer, as they are composed of two layers of phosphate-containing lipids with proteins floating between these layers. This forms through the process of “self assembly” by which the hydrophobic tails of phospholipid molecules secure an inward position within the membrane. This prevents it from contacting water on a normal basis. Conversely, the hydrophilic heads of the phospholipids are positioned outward and are in contact with water. The proteins in the structure are responsible for carrying out the majority of the functions specific to the membrane and imparts the selectivity to certain materials that can pass through the membrane.
Figure 2: A phospholipid bilayer (cell membrane) with embedded proteins.
Cell Wall
Many cells within the prokaryotic and eukaryotic families also have cell walls outside the cell membrane that help to protect them and provide strength and support; although, animal cells and protozoa do not have cell walls. Unlike the cell membrane, the cell wall is not selective and does not allow materials to pass through easily. Prokaryotic cells have a thick, rigid cell wall composed of amino acids and sugars (peptidoglycan), but the cell wall composition within eukaryotes varies. For example, fungi cell walls include a polysaccharide called chitin while plants exhibit cell walls with the polysaccharide cellulose.
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Cell Structure and Function
Prokaryotes and eukaryotes must also regulate nutrients and wastes, and require a supply of energy to exist.
Metabolic activities such as photosynthesis and respiration can be carried out by both cell types. However, photosynthetic activity initiates in the chloroplasts in eukaryotes, while it occurs in the thylakoid in prokaryotes. The thylakoid is an extension of the plasma membrane that contains photosynthetic pigments.
Prokaryotic Characteristics
Pili
Both physical and chemical diversity can be viewed between prokaryotic and eukaryotic cells. A structural examination can begin by reviewing major prokaryotic structures, such as the flagella, although prokaryotes tend to have fewer organelles and diverse features than eukaryotes. Flagella are long, cylindrical protrusions on the outside of a cell. They are often, but not always, present, and facilitate mobility through a whiplike, motor rotation. For example, E. coli bacteria rotate their flagellum in a clockwise direction to swim through
Cell Wall their environment.
Ribosomes
Nucleoid
Region
Some prokaryotic cells also have a glycocalyx, which Cytoplasmic is a slime-coating used to protect the cell. The gly- Membrane cocalyx also allows prokaryotes to attach to different surfaces (such as teeth, lungs, or even artificial joints).
This is helpful as these surfaces provide a nutritional resource and often a safe habitat.
Cytoplasm
Many prokaryotic bacteria cells also posses pili. Pili are small, hair-like structures on the outer surface of many
Flagella
types of bacteria that provide the ability to adhere to hosts and other bacteria, as well as the ability to trans- Figure 3: Prokaryotic cell diagram showing some of mit genetic information. Bacterial cells that posses sex the major structures including flagella, pili, freely pili have the ability to transmit genetic information with existing DNA, and ribosomes. other cells. This may result in the transmission of genes that provide antibacterial (antibiotic) resistance.
Eukaryotic Characteristics
Eukaryotic cells are generally more structurally complex than prokaryotic cells and typically contain more components. A network of membrane-bound organelles exists within each eukaryotic cell. Essentially, these
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Cell Structure and Function organelles function as “mini” organs within the cell. Each organelle fulfills a
Did You Know... unique function, but they must all work together to maintain homeostasis within the cell.
Red blood cells are created in
?
Nucleus
The nucleus is often referred to as the control center of the cell. It contains nuclear DNA, which codes for the majority of the proteins required for human survival. The nucleus is surrounded by a double membrane system, called the nuclear envelope. This structure contains nuclear pores that facilitate communication between the nucleus and other cellular structures via active and passive transport of molecules and ions. A smaller, dense structure called the nucleolus also exists within the nucleus. The nucleolus is composed of RNA and proteins. This structure functions to create the ribosomal
RNA (rRNA) and assemble necessary ribosomes within the cell. The exact size of this structure is dependent upon the necessary amount of ribosomes.
For example, if great need for ribosomes exists within the cell, the nucleolus is typically larger. Conversely, the nucleolus remains smaller if only a small number of ribosomes is required.
the bone marrow. Once they mature and enter the blood stream their nucleus degrades and they live the rest of their lives without one.
This frees up space to carry oxygen and makes it easier to fit through small capillaries.
The Mitochondrion
While most eukaryotic DNA is housed within the nucleus, mitochondrial DNA is located in a cell organelle called the mitochondrion. Mitochondria (more than one mitochondrion) are separate organelles located in the cytoplasm, and are often considered the powerhouse of the cell because of their functional involvement in energy metabolism. In fact, this small, elongated structure is the site for cellular respiration in which nearly all of the energy of the cell is produced (when oxygen is present). Mitochondrial DNA enFigure 4: Mitochondrion. codes genes which aid in energy production. Mitochondria are housed within a double membrane structure. The outer membrane serves to hold and protect the organelle while the inner membrane, called the cristae, contains multiple folds that allow for a greater surface area.
The increased surface area allows for more efficient energy production. There is always at least one mitochondrion in eukaryotic cells. However, cells which require a great amount of energy, such as muscle cells, may contain thousands of individual mitochondria.
The Cytomembrane System
Within the entire network of the cell lies the cytomembrane system composed of the endoplasmic reticulum, golgi body and many vesicles. This system is responsible for the production of lipids and proteins. The
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Cell Structure and Function cytomembrane system begins in the endoplasmic reticulum (ER) which, in animal cells, is connected directly to the outer layer of the nuclear envelope. The
ER is divided into two components, the rough ER and the smooth ER. The rough ER is composed of flattened sac-like structures with attached ribosomes and functions in protein synthesis and protein transportation to the golgi body. The term “rough ER” is derived from this rough appearance. The surface appears bumpy or rough due to the ribosomes which are attached to it. The smooth ER is also composed of flattened sac-like structures, and is devoid of ribosomes.
Thus, the smooth ER appears smooth under a microscope. The smooth ER functions to create many of the lipids of the cell and also functions in detoxification of destructive chemicals that may pass throughout the cell. 2
1
3
Figure 5: The cytomembrane system. Note the endoplasmic reticulum (1) sprouting from the outer layer of the nuclear envelope (2), and the flattened, saclike appearance of the golgi apparatus (3) in the lower left.
The next step in the cytomembrane system is the Golgi body, also called the Golgi apparatus. This structure is composed of flattened sac-like structures stacked on top of each other. The proteins and lipids created in the ER are transferred to the Golgi body where they are modified, packed and then shipped out to specified locations both within and outside of the cell.
After the proteins and lipids are modified and packed in the Golgi body, they are then transported in vesicles. One type of vesicle, called the secretory vesicle, is composed of a sac freed or pinched off of the Golgi body. Secretory vesicles transport proteins and lipids to the plasma membrane so they can be released or secreted from the cell.
Another type of vesicle, the lysosome, is also released off the Golgi body. These vesicles contains enzymes that are used in the digestion of cellular components. They can digest proteins, lipids, carbohydrates, cell parts and even an entire cell.
Another membrane bound vesicle is the peroxisome. The peroxisome is responsible for lipid and amino acid break down, as well as the break down of hydrogen peroxide, a highly toxic byproduct of certain cellular reactions. When drinking alcohol, peroxisomes also function to break down almost half of all that was consumed.
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Cell Structure and Function
The Cytoskeleton
The cytoskeleton is a network of interconnected fibers within the cell that provides shape, internal organization, and the ability for a cell to move. Microtubules are a part of the cytoskeleton of a cell. These hollow cylinders are composed of tubulin protein and have a diameter of about 25 nm. Microtubules are key in cellular structure, providing support in the cytoplasm. They are also very important in cellular division as they form the centrioles and spindle fibers necessary for the cells to divide. Microtubules can also function in the movement of the cell by combining to form flagella and cilia. Flagella are long, cylindrical protrusions on the outside of a cell. They provide mobility through a whip-like rotation. Cilia are much smaller, hair-like and typically more abundant than flagella. The cilia beat together to either allow the cell to move or to cause movement of extracellular fluid.
The side kick to the microtubules are the microfilaments, also known as actin filaments. Microfilaments are much smaller and thinner than their microtubule counterpart, but are widely abundant throughout the cell.
They are made of the protein actin and are typically no larger than 8 nm (Remember: 1 nanometer (nm) = 1 x 10-9 meter) in diameter. These cellular components are also responsible for cellular structure within the cytoplasm along with cellular movement. Another support component within the cytoplasm is the intermediate filaments. These filaments are about 10 nm in diameter and provide additional strength to the cytoplasm of the cell. There are many cellular components that work together to facilitate all the responsibilities of a cell.
Each individual component works to perform its specific function so that the cell, as a whole, can perform the major functions required of it.
Figure 6: Parts of the cytoskeleton.
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Cell Structure and Function
Animal Cell (top); Plant Cell (bottom)
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Figure 7: Animal and plant cells are both eukaryotic.
Cell Structure and Function
Quick Reference: Eukaryotic Organelles
•
Cytoskeleton: The cell “skeleton” found in all eukaryotic cells that provides shape to the cell while also enabling it to move.
•
Endoplasmic Reticulum (ER): A series of membranes extending throughout the cytoplasm that can be peppered with ribosomes (rough ER) or not (smooth ER). The rough ER functions in protein synthesis while the smooth ER functions in lipid synthesis.
•
Golgi Apparatus (also called the Golgi Body): A series of flattened sac-like bodies that processes the cell’s proteins and lipids before they are released to their final destination.
•
Lysosomes: Enzyme-filled vesicles found within the cell that aid in intracellular digestion.
•
Mitochondrion (plural: mitochondria): The “power plant” of the cell. These are membrane bound organelles (inner and outer membrane) with their own circular DNA, and make ATP (energy) for the rest of the cell.
•
Nuclear Envelope: An outer membrane that surrounds the nucleus.
•
Nuclear Pores: Holes in the nuclear envelope that permit communication between the internal nuclear environment and the cytoplasm.
•
Nucleus: Houses the genetic content (DNA) of the cell.
•
Nucleolus (plural: nucleoli): A part of the nucleus that is made of RNA, protein and chromatin; and, manufactures rRNA and ribosomes.
•
Peroxisomes: Contain enzymes that help the cell destroy toxins.
•
Ribosomes: Ribosomes are large molecules found in all living cells. Ribosomes the site of protein synthesis. A strand of mRNA docks onto a ribosome. The correct amino acids are then recruited to the ribosome to create a protein.
•
Vacuole: A fluid-filled organelle that helps isolate its contents from the cytoplasm.
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Cell Structure and Function
Table 1: Structural Variations Between Prokaryotic and Eukaryotic Cells
Structure
Present in Prokaryotic Cell?
Present in Eukaryotic Cell?
Nucleus
No
Yes
Plasma Membrane
Yes
Yes
Cell Wall
Yes
Yes (plant and fungal cells; not animal cells)
Cytoplasm
Yes
Yes
Flagella
Occasionally
Occasionally
Pili
Occasionally
No
Cilia
No
Occasionally
Glycocalyx
Occasionally
Occasionally
Cytoskeleton
No
Yes
Endoplasmic Reticulum
No
Yes
Mitochondria
No
Yes
Golgi Apparatus
No
Yes
Chloroplast
No
Yes, in plants and other photosynthetic organisms
Ribosome
Yes
Yes
Lysosome
No
Yes
Peroxisome
No
Yes
Vacuole and Vesicle
Occasionally
Yes (in most cells)
Pre-Lab Questions
1. Identify three major similarities and differences between prokaryotic and eukaryotic cells.
2. Where is the DNA housed in a prokaryotic cell? Where is it housed in a eukaryotic cell?
3. Identify three structures which provide support and protection in a eukaryotic cell.
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Cell Structure and Function
Experiment 1: Identifying Cell Structures
The structure of a cell dictates the majority of its function. You will view a selection of slides that exhibit unique structures that contribute to tissues function.
Materials
Onion Root (allium) Digital Slide Images
Procedure
1. Examine the onion root tip digital slide images on the following pages. Then, respond to the Post-Lab
Questions.
Onion Root Tip: 100X
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Cell Structure and Function
Cytoplasm
Chromosomes
Chromosomes
Cell wall
Nucleus
Onion Root Tip: 1000X
Cytoplasm
Cell wall
Nucleus
Chromosomes
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Onion Root Tip: 1000X
Cell Structure and Function
Onion Root Tip: 100X. Each dark circle indicates a different nucleus.
Cell wall
Cytoplasm
Chromosomes
Onion Root Tip: 1000X
Nucleus
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Cell Structure and Function
Post-Lab Questions
1. Label each of the arrows in the following slide image:
A
C
D
Onion Root Tip: 1000X
B
2. What is the difference between the rough and smooth endoplasmic reticulum?
3. Would an animal cell be able to survive without mitochondria? Why or why not?
4. What could you determine about a specimen if you observed a slide image showing the specimen with a cell wall, but no nucleus or mitochondria?
5. Hypothesize why parts of a plant, such as the leaves, are green, but other parts, such as the roots, are not. Use scientific reasoning to support your hypothesis.
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Cell Structure and Function
Experiment 2: Create a Cell
In this experiment, you will create an animal and a plant cell using household items to observe the difference between prokaryotes and eukaryotes.
Materials
30 cm x 30 cm Aluminum Foil (Cell Wall)
2 Resealable Bags (Cell Membrane)
20 Beads, any color (Rough Component of Endo- *Bowl plasmic Reticulum and free ribosomes)
*Household items to represent the Golgi bodies
2 Eggs (plastic) (Nucleus)
and chloroplasts
4 Gelatin Packets, unflavored
*Warm Water
12 Kidney Beans (Mitochondria)
Permanent Marker
*You Must Provide
4 Pipe Cleaners (Endoplasmic Reticulum)
Procedure
1. Place four packets of unflavored gelatin in a bowl. Add 4 cups of hot water to the bowl. Do not refrigerate the mixture yet!
Note: You do not need to heat the water in a microwave. Simply run tap water until it feels warm to the touch.
2. Label each resealable bag as either “Plant Cell” or “Animal Cell”. These will serve as the cell membrane.
3. Construct rough endoplasmic reticulum by stringing 10 beads onto two of the pipe cleaners (5 on each). The remaining, unbeaded pipe cleaners represent smooth endoplasmic reticulum.
4. Construct a cell wall using the aluminum foil. This should be large enough to fit the resealable bag when filled with half of the gelatin and some of the cell structures.
Hint: It is helpful to make this square-shaped.
5. Using your knowledge of the Golgi apparatus and chloroplasts, think of two household items which can represent these structures. Find and collect these items for use in this experiment.
Hint: Colored paper may bleed when placed in gelatin.
6. Open the resealable bag labeled “Plant Cell” and pour half of the liquid gelatin into it.
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Cell Structure and Function
7. Add the items which represent plant cell structures (you must determine which items!) into the gelatin and tightly close the bag. If there is an “organelle” present in both plant and animal cells make sure to leave enough to be included in the animal cell.
8. Place the bag in the aluminum foil cell wall.
9. Open the resealable bag labeled “Animal Cell” and pour the remainder of the gelatin into it.
10. Add the items which represent animal cell structures (you must determine which items!) into the gelatin and tightly close the bag.
11. Place both “cells” into the refrigerator for 24 hours.
12. Return after 24 hours and observe the “cells” you have made. Notice the difference between the animal cell and the plant cell.
Post-Lab Questions
1. What cell structures did you place in the plant cell that you did not place in the animal cell?
2. Is there any difference in the structure of the two cells?
3. What structures do organisms that lack cell walls have for support?
4. How are organelles in a cell like organs in a human body?
5. How does the structure of a cell suggest its function? List three examples.
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Lab 11
Mitosis
Mitosis
Learning Objectives
•
Relate DNA to genes, alleles and chromosomes
•
Identify chromosomal structure during the various stages of the cell cycle including chromatin, sister chromatids, homologous chromosomes, and centromeres
•
Compare and contrast animal and plant mitosis
•
Apply the concept of ploidy to the human species
•
Explain the role of parent and daughter cells in cell cycle division
Introduction
All of a cell’s hereditary information is encoded by the genome. The genome, all of an organism’s genes, is vital for life as it directs the formation of all the proteins the cell needs to survive. It also allows individual cells to work together and form complex, multicellular organisms. However, both unicellular and multicellular organisms must be able to grow and divide for life to continue. Cell cycle division provides a mechanism by which a cell can pass along its DNA from one generation to the next, and ensure species propagation. Mitosis is the term for eukaryotic cell division, while binary fission describes prokaryotic division.
DNA and Genes
In eukaryotes, the genome is encoded by chromatin, which is composed of deoxyribonucleic acid (DNA), ribonucleic acid
(RNA), and proteins. DNA is a double-stranded molecule, constructed from repeating units of a phosphate molecule, a sugar (deoxyribose), and one of four possible nucleic acids adenine, thymine, cytosine, or guanine. Units which incorporates these three structures are called nucleotides. Hydrogen bonds between complementary nucleotides (adenine and thymine [A - T] and cytosine to guanine [C - G]) hold DNA strands together. Sections of DNA that code for traits such as eye color are called genes (Figure 1).
Ploidy
Figure 1: Chromosomes are long strands of
DNA coiled up. Pieces of DNA that code for traits are called genes.
The number of genes required for an organism to survive varies by species. In fact, different species may have a different number of chromosomes, as well as a different number of copies of each chromosome. Somatic (any body cells, but sex cells) human cells have two copies of 23 chromosomes, resulting in 46 chromosomes per cell. These cells are considered diploid, because they have two copies.
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Mitosis
Almost all somatic, mammalian cells have two complete sets of chromosomes. One set originates from the female parent and one set originates from the male parent, resulting in pairs of homologous chromosomes
(Figure 2).
Cell Cycle Division
Cell cycle division occurs when a parent cell copies its entire genome and splits into two identical daughter cells. Do not confuse the word
“parent” with mother or father. It is just a term to differentiate the cell that is splitting from the two new cell created. Each new daughter cell (not to
Figure 2: Sister chromatids (a) and be associate with the female sex) contains two complete sets of chromohomologous chromosomes (b) of somes. The cellular content is identical to the parent cell as long as no a diploid organism in which 2n=4. mutations or errors occur during cell cycle division.
Cell cycle division is divided into three phases: interphase, mitosis, and cytokinesis (Figure 3). These phases are further broken down into individual stages. These stages are identified as:
Interphase; Stage:
•
Gap 1 (G1): The first stage of interphase; or, the first growth phase.
•
S: DNA synthesis. The second stage of interphase.
•
Gap 2 (G2): The third stage of interphase; or, the second growth stage.
Mitosis; Stage:
•
Prophase
•
Metaphase
•
Anaphase
•
Telophase
Cytokinesis:
•
Figure 3: Mitosis is only one stage of the cell cycle.
There are no further subdivided stages within cytokinesis.
A cell normally completes the cycle in 18 - 24 hours. Mitosis only occupies one to two hours of that period.
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Mitosis
Each stage is regulated by specialized proteins, such as cyclins and cyclin-dependent kinases (CDKs) that coordinate the division and cell growth. Certain types of cancer are associated with the failure of these proteins.
During interphase, the cell prepares for mitosis and cytokinesis by growing larger in size, duplicating its internal organelles (G1), replicating its DNA (S) and preparing for cell division (G2).
You can easily remember the activity which occurs during the S phase because it is when new DNA is synthesized (synthesized starts with s). When the full set of DNA is replicated, two identical copies of each chromosome exist in the parent cell. These identical copies pair up and are referred to as sister chromatids. Sister chromatids exist as loose strands of genetic content known as chromatin during interphase.
These structures don’t condense into chromosomes until prophase of mitosis. To form a chromosome, two sister chromatids are joined together as a four-arm structure by a centromere (see prophase stage below).
Chromosomes also incorporate a variety of proteins, such as histones, which tightly package the DNA. Interestingly, if the chromatin from a single human cell was uncoiled and stretched out, it would be over two meters long. However, it is important that it remain in a loose configuration to permit transcription of DNA.
After the DNA is fully replicated, the cell engages in “proofreading” and additional protein production (G2). In this process, proteins involved in DNA replication (such as DNA polymerase) review the replicated content and correct any errors which may have been made during replication. G1 and G2 are also responsible for initiating a variety of checkpoints which verify that the size and environmental growth factors are sufficient prior to entering mitotic division.
Mitosis follows interphase. It is during mitosis that sister chromatids segregate into two daughter cells. The four stages of mitosis are described below:
Mitosis (Figure 2):
•
Prophase: Loosely formed chromatin condenses into tightly packed chromosomes. Centromeres attach themselves to coiled (condensed) sister chromatids to hold their structure together.
The nuclear envelope of the parent cell breaks down. Pairs of centrioles, which serve as cellular anchors during division, appear at the mitotic spindle, located at the cellular poles at separate sides of the cell.
•
Metaphase: Sister chromatids migrate towards the metaphase plate. Microtubules (long strands) grow from the centrioles and link together while attaching to each pair of sister chroma-
163
Mitosis tids. The orientation of each pair of sister chromatids is independent from all other pairs. This means they can “flip flop” as they line up, effectively shuffling their genetic information into new combinations. •
Anaphase: The microtubules attached to the centromeres shorten This causes the sister chromatids to separate and move toward opposite poles of the parent cell. At this point, each sister chromatid is now an individual chromosome. Microtubules attached to other microtubules from the opposite pole lengthen, effectively elongating the cell.
•
Telophase: One set of chromosomes arrives at each pole. A new nuclear envelope begins to form, chromosomes are uncoiled back into chromatin, and a new nucleus is formed at each end of the cell.
Cell cycle division concludes with cytokinesis. During animal cell cytokinesis, the plasma membrane of the cell folds in and encloses around each nucleus, creating two, diploid daughter cells.
Mitosis is virtually identical in plant and animal cells, however, there is one small difference which occurs during telophase. Plants, due to the presence of a cell wall, cannot “pinch” the cytoplasm into two daughter cells. Instead, a new cell wall must be developed which will then separate the two cells, allowing them both to be fully covered with a cell wall.
Prophase
Metaphase
Anaphase
Telophase
Figure 4: The stages of mitosis.
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Cytokinesis
Mitosis
Definitions
Alleles – Alternate forms of the same gene.
Centriole – An organelle that forms the microtubules which separate chromosomes during cell division. Centromere – The region of the chromosome where chromatids are held together. This forms the kinetochore. This is also where microtubules attach during cell division.
Chromatids – One of two identical chromosomes (often called “sister chromatids”).
Chromosome – A threadlike structure consisting of chromatin that contains a long, continuous strand of DNA and its associated proteins.
Cytokinesis – The final step of the cell cycle where the cell splits to produce two daughter cells.
Diploid – Cells that have two copies (2n) of each chromosome.
Gene – A sequence of DNA that codes for a specific detectable product such as a protein or
RNA.
Genetic Variation – The diversity in the DNA found in otherwise similar organisms.
Homologous Chromosome – Chromosomes that contain the same genes, but potentially different alleles.
Locus – The exact position of a gene on a chromosome.
Microtubules – Long polymer strands of protein that separate chromosomes during cell division.
Ploidy – The number (n) of copies of each chromosome contained in a cell (usually 1n or 2n in animals). Tetrad – Four homologous chromosomes, two of each pair of sister chromatids, that line up during prophase I in meiosis.
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Mitosis
Pre-Lab Questions
1. What are chromosomes made of?
2. Research the differences that exist between mitosis and binary fission. Identify at least one difference, and explain why it is significant.
3. Cancer is a disease related to uncontrolled cell division. Investigate two known causes for these rapidly dividing cells and use this knowledge to invent a drug that would inhibit the growth of cancer cells.
Experiment 1: Observation of Mitosis in a Plant Cell
In this experiment, we will look at the different stage of mitosis in an onion cell. Remember that mitosis only occupies one to two hours while interphase can take anywhere from 18 - 24 hours. Using this information and the data from your experiment, you can estimate the percentage of cells in each stage of the cell cycle.
Materials
Onion (allium) Root Tip Digital Slide Images
Procedure
Part 1: Calculating Time Spent in Each Cell Cycle Phase
1. The length of the cell cycle in the onion root tip is about 24 hours. Predict how many hours of the 24 hour cell cycle you think each step takes. Record your predictions, along with supporting evidence, in
Table 1.
2. Examine the onion root tip slide images on the following pages. There are four images, each displaying a different field of view. Pick one of the images, and count the number of cells in each stage. Then count the total number of cells in the image. Record the image you selected and your counts in Table 2.
3. Calculate the time spent by a cell in each stage based on the 24 hour cycle:
Hours of Stage = 24 x Number of Cells in Stage
Total Number of Cells Counted
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Mitosis
Part 2: Identifying Stages of the Cell Cycle
1. Observe the images of the root cap tip.
2. Locate a good example of a cell in each of the following stages: interphase, prophase, metaphase, anaphase, and telophase.
3. Draw the dividing cell in the appropriate area for each stage of the cell cycle, exactly as it appears. Include your drawings in Table 3.
Nucleus
Cytoplasm
Chromosomes
Cell wall
Onion Root Tip: 100X
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Mitosis
Onion Root Tip: 100X
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Onion Root Tip: 100X
Mitosis
Onion Root Tip: 100X
Table 1: Mitosis Predictions
Predictions:
Supporting Evidence:
Table 2: Mitosis Data
Number of Cells in Each Stage
Total Number of Cells
Calculated % of Time Spent in Each Stage
Interphase:
Interphase:
Prophase:
Prophase:
Metaphase:
Metaphase:
Anaphase:
Anaphase:
Telophase:
Telophase:
Cytokinesis:
Cytokinesis:
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Mitosis
Table 3: Stage Drawings
Cell Stage:
Interphase:
Prophase:
Metaphase:
Anaphase:
Telophase:
Cytokinesis:
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Drawing:
Mitosis
Post-Lab Questions
1. Label the arrows in the slide image below with the appropriate stage of the cell cycle.
A
C
B
F
E
D
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Mitosis
2. In what stage were most of the onion root tip cells? Based on what you know about cell cycle division, what does this imply about the life span of a cell?
3. Were there any stages of the cell cycle that you did not observe? How can you explain this using evidence from the cell cycle?
4. As a cell grows, what happens to its surface area to volume ratio? (Hint: Think of a balloon being blown up). How does this ratio change with respect to cell division?
5. What is the function of mitosis in a cell that is about to divide?
6. What would happen if mitosis were uncontrolled?
7. How accurate were your time predication for each stage of the cell cycle?
8. Discuss one observation that you found interesting while looking at the onion root tip cells.
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Mitosis
Experiment 2: Tracking Chromosomal DNA Movement through Mitosis
Although mitosis and meiosis share similarities, they are different processes and create very different results.
In this experiment, you will follow the movement of the chromosomes through mitosis to create somatic daughter cells.
Materials
2 Sets of Different Colored Pop-it® Beads (32 of each - these may be any color)
(8) 5-Holed Pop-it® Beads (used as centromeres)
Procedure
Genetic content is replicated during interphase. DNA exists as loose molecular strands called chromatin; it has not condensed to form chromosomes yet.
Sister chromatids begin coiling into chromosomes during prophase. Begin your experiment here:
1. Build a pair of replicated, homologous chromosomes. 10 beads should be used to create each individual sister chromatid (20 beads per chromosome pair). Two five-holed beads represent each centromere. To do this...
a. Start with 20 beads of one color to create your first sister chromatid pair. Five beads must be snapped together for each of the four different strands. Two strands create the first chromatid, and two strands create the second chromatid.
b. Place one five-holed bead flat on a work surface with the node positioned up. Then, two of the four strands into the bead to create an “I” shaped sister
Figure 5: Bead set-up. The blue beads reprechromatid. Repeat this step with the other two sent one pair of sister chromatids and the strands and another five-holed bead. black beads represent a second pair of sister
c. Once both sister chromatids are constructed, con- chromatids. The black and blue pair are homologous. nect them by their five-holed beads creating an “X” shape. d. Repeat this process using 20 new beads (of a different color) to create the second sister chromatid pair. See Figure 5 for reference.
2. Assemble a second pair of replicated sister chromatids; this time using 12 beads, instead of 20, per pair
(six beads per each complete sister chromatid strand).
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Mitosis
3. Repeat this process using 12 new beads (of a different color) to create the second set of sister chromatids. See Figure 6 for reference.
4. Configure the chromosomes as they would appear in each of the stages of the cell cycle (prophase, metaphase, anaphase, telophase, and cytokinesis). Diagram the images for each stage in the section titled “Cell Cycle Division: Mitosis
Beads Diagram”. Be sure to indicate the number of chromoFigure 6: Second set of replicated chromosomes present in each cell for each phase. somes. Cell Cycle Division: Mitosis Beads Diagram:
Prophase
Metaphase
Anaphase
Telophase
Cytokinesis
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Mitosis
Post-Lab Questions
1. How many chromosomes did each of your daughter cells contain?
2. Why is it important for each daughter cell to contain information identical to the parent cell?
3. How often do human skin cells divide? Why might that be? Compare this rate to how frequently human neurons divide. What do you notice?
4. Hypothesize what would happen if the sister chromatids did not split equally during anaphase of mitosis.
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Mitosis
Experiment 3: The Importance of Cell Cycle Control
Some environmental factors can cause genetic mutations which result in a lack of proper cell cycle control
(mitosis). When this happens, the possibility for uncontrolled cell growth occurs. In some instances, uncontrolled growth can lead to tumors, which are often associated with cancer, or other biological diseases.
In this experiment, you will review some of the karyotypic differences which can be observed when comparing normal, controlled cell growth and abnormal, uncontrolled cell growth. A karyotype is an image of the complete set of diploid chromosomes in a single cell.
Materials
*Computer Access
*Internet Access
*You Must Provide
Procedure:
1. Begin by constructing a hypothesis to explain what differences you might observe when comparing the karyotypes of human cells which experience normal cell cycle control versus cancerous cells (which experience abnormal, or a lack of, cell cycle control). Record your hypothesis in Post-Lab Question 1.
Note: Be sure to include what you expect to observe, and why you think you will observe these features. Think about what you know about cancerous cell growth to help construct this information
2. Go online to find some images of abnormal karyotypes, and normal karyotypes. The best results will come from search terms such as “abnormal karyotype”, “HeLa cells”, “normal karyotype”, “abnormal chromosomes”, etc. Be sure to use dependable resources which have been peer-reviewed
3. Identify at least five abnormalities in the abnormal images. Then, list and draw each image in the Data section at the end of this experiment. Do these abnormalities agree with your original hypothesis?
Hint: It may be helpful to count the number of chromosomes, count the number of pairs, compare the sizes of homologous chromosomes, look for any missing or additional genetic markers/flags, etc.
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Mitosis
Data
1.
2.
3.
4.
5.
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Mitosis
Post-Lab Questions
1. Record your hypothesis from Step 1 in the Procedure section here.
2. What do your results indicate about cell cycle control?
3. Suppose a person developed a mutation in a somatic cell which diminishes the performance of the body’s natural cell cycle control proteins. This mutation resulted in cancer, but was effectively treated with a cocktail of cancer-fighting techniques. Is it possible for this person’s future children to inherit this cancer-causing mutation? Be specific when you explain why or why not.
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Lab 12
Meiosis
Meiosis
Learning Objectives
•
Compare and contrast gametes and somatic cells including haploid and diploid
•
Explain how independent assortment, crossing over and mutations influence genetic variation
•
Apply meiosis to the development of genetic diseases
Introduction
Meiosis, also known as gametogenesis, only occurs in sexually reproducing organisms (plants and animals). In humans, meiosis generates four haploid cells in both males and females. These cells are called gametes. Male gametes are referred to as sperm cells, while female gametes are referred to as egg cells. Haploid cells only contain one copy (1n) of chromosomal content (n = 23 chromosomes in humans). Figure 1: Haploid gametes fuse during fertilization to form a diploid cell.
Haploid gametes fuse during fertilization and form a diploid cell. The newly fused cell contains two copies of each chromosome (2n = 46 chromosomes in humans); these chromosomes are now referred to as homologous chromosomes. This is unlike somatic cells which are produced through mitosis and already contain two copies of chromosomal content .
Meiosis I
Meiosis is similar to mitosis, but involves a second round of cellular division, therefore producing four daughter cells instead of two. The two rounds of cell division are called meiosis one and meiosis two. Meiosis I produces two haploid cells from the primary gamete.
The stages of meiosis I mirror the stages you learned about in mitosis except the two new cells are haploid instead of diploid. This includes prophase, metaphase, anaphase and telophase. However, because there are two rounds of meiosis, each stage also receives a “1” or a “2” after the name, depending on which round of division the cell is currently in. These stages are also indicated with Roman Numerals. You may recall that mitosis is preceded by a period called interphase during which cellular content is duplicated and DNA is proofread for accuracy. The same is true for meiosis, as an interphase period prepares the cell for successful meiotic division.
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Meiosis
The stages of meiosis I are outlined as follows:
•
Prophase I: DNA condenses into chromosomes. Centromeres attach themselves to coiled
(condensed) sister chromatids to hold their structure together. Homologous chromosomes pair up and form a chiasma. Crossing over may occur at this point. This means that homologous chromosomes may exchange and recombine regions of DNA. Crossing over creates a unique genetic fingerprint for the daughter cells, and contributes to the diversity seen in progeny.
Similar to mitotic prophase, the nuclear envelope of the parent cell breaks down in this phase as well. Pairs of centrioles, which serve as cellular anchors during division, appear at the mitotic spindle, located at the cellular poles.
•
Metaphase I: Homologous chromosomes migrate towards the metaphase plate. Microtubules grow from the centrioles and either link together or attach to each pair of homologous chromosomes at the centromeres. The orientation of each pair of sister chromatids is independent from all other pairs. This action is referred to at the Law of Independent Assortment. This means the chromosomes can “flip flop” as they line up, effectively shuffling their genetic information into new combinations, providing a second source of genetic variation.
•
Anaphase I: Microtubules attached to chromosomes. They then shorten in length, which pulls the homologous chromosomes apart (sister chromatids remain paired) and moving them toward opposite poles. Microtubules attached to other microtubules from opposite poles lengthen to elongate the cell.
•
Telophase I: One set of sister chromatids arrives at each pole. A new nuclear envelope begins to form, chromosomes are uncoiled back into chromatin, and a new nucleus is formed at each end of the cell.
Cytokinesis follows meiosis one, deepening the cellular cleavage formed in telophase one and fully dividing the two new daughter cells by engulfing each nuclei within the plasma membrane.
It is important to note that unlike mitosis, meiosis I creates haploid (not diploid) cells. This is because the homologous chromosomes during the first found of meiosis. They do not divide into the same cells. Instead, the sister chromatids remain coupled and are partitioned into the cell products. Therefore, the two daughter cells produced from meiosis one contain identical genetic content. They only contain one copy of the genome because all of the alleles are the same.
Meiosis Two
The two daughter cells produced during meiosis one immediately enter into prophase two of meiosis II. The meiosis two stages are described as follows:
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Meiosis
•
Prophase II: New spindle fibers form as the nucleus break down again. Chromosomes condense in the form of sister chromatids.
•
Metaphase II: Sister chromatids migrate towards the metaphase plate. Similar to metaphase one, microtubules attach to the centromeres.
•
Anaphase II: This time, sister chromatids (rather than homologous chromosomes) are separated as the microtubules shorten and pull them apart. The cell elongates as microtubules attached to additional microtubules from opposing poles lengthen.
•
Telophase II: 23 chromatids arrive at each pole. A new nucleus forms around each.
Cytokinesis again follows, resulting in four gametes.
Meiosis I
Prophase I
Metaphase I
Anaphase I
Telophase I
Meiosis II
Prophase II
Metaphase II
Anaphase II
Telophase II
Figure 2: Stages of Meiosis I and II.
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Meiosis
Checkpoints and Mutations
Meiotic division is complex and highly regulated. There are a series of checkpoints that a cell must pass before the next stage of division will begin. When these checkpoints function properly, mutations and errors are identified and repaired. Some of these checkpoints include verifying that ample nutrients are available for division and proofreading replicated
DNA strands. Problems which are found may result in a pause within meiosis to allow the appropriate proteins sufficient time to repair the error. If the error can’t be repaired, the cell may attempt apoptosis (cell death).
Checkpoints that are missing or do not function correctly may result in genetic mutations. Genetic mutations which occur in somatic cells are not heritable mutations. This means that the mutation will not be propagated amongst progeny. Mutations which occur in gametes are heritable mutations which risk genetic expression amongst future generations.
?
Did You Know...
There are many types of mutations. These range in scale from silent to lethal, and may affect a single nucleotide or a full chromosome. Trisomy 21, or Down’s Syndrome, is one example of a chromosomal abnormality. In this disease, three copies of chromosome
21 exist rather than the standard two copies. This type of abnormality is typically lethal, however, chromosome 21 is much smaller than many other chromosomes and thus the progeny are viable.
One mutation of particular concern is when the amount of genetic material existing in a cell changes. It is critical that a gamete contains only one copy of each chromosome found in a parent cell.
Definitions (Review):
Alleles – Alternate forms of the same gene.
Centriole – An organelle that forms the microtubules which separate chromosomes during cell division.
Centromere – The region of the chromosome where chromatids are held together. This forms the kinetochore. This is also where microtubules attach during cell division.
Chiasma – The point where crossover occurs in meiosis one.
Chromatids – One of two identical chromosomes (often called “sister chromatids”).
Chromosome – A threadlike structure consisting of chromatin that contains a long, continuous strand of DNA and its associated proteins.
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Meiosis
Crossing Over – The process, during meiosis I, by which two, paired homologous chromosomes are paired and exchange regions of DNA.
Cytokinesis – The final step of the cell cycle where the cell splits to produce two daughter cells.
Diploid – Cells that have two copies (2n) of each chromosome.
Haploid – Cells that have one copy (1n) of each chromosome.
Gamete – A cell containing a single copy of each chromosome (egg cells and sperm cells).
Gene – A sequence of DNA that codes for a specific detectable product such as a protein or RNA.
Genetic Variation – The diversity in the DNA found in otherwise similar organisms.
Homologous Chromosome – Chromosomes that contain the same genes, but potentially different alleles. Locus – The exact position of a gene on a chromosome.
Meiosis – The stage of the cell cycle when the nucleus is divided into daughter cells (gametes: sperm cells and egg cells) with a reduced number of chromosomes (from diploid to haploid).
Microtubules – Long polymer strands of protein that separate chromosomes during cell division.
Ploidy – The number (n) of copies of each chromosome contained in a cell (usually 1n or 2n in animals).
Tetrad – Four homologous chromosomes, two of each pair of sister chromatids, that line up during prophase I in meiosis.
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Meiosis
Pre-Lab Questions
1. Compare and contrast mitosis and meiosis.
2. What major event occurs during interphase?
Experiment 1: Following Chromosomal DNA Movement through Meiosis
In this experiment, you will model the movement of the chromosomes through meiosis I and II to create gametes.
Materials
2 Sets of Different Colored Pop-it® Beads (32 of each - these may be any color)
(8) 5-Holed Pop-it® Beads (used as centromeres)
Procedure
Part 1: Modeling Meiosis without Crossing Over
As prophase I begins, the replicated chromosomes coil and condense…
1. Build a pair of replicated, homologous chromosomes. 10 beads should be used to create each individual sister chromatid (20 beads per chromosome pair). The five-holed beads represent each centromere. To do this...
a. Start with 20 beads of the same color to create your first sister chromatid pair. Five beads must be snapped together for each of the four different strands. Two strands create the first chromatid, and two strands create the second chromatid with a 5holed bead at the center of each chromatid. This creates an “I” shape.
b. Connect the “I” shaped sister chromatids by the 5- Figure 3: Bead set-up. The blue beads represent one pair of sister chromatids and the holed beads to create an “X” shape. black beads represent a second pair of sister
c. Repeat this process using 20 new beads (of a differ- chromatids. The black and blue pair are hoent color) to create the second sister chromatid pair. mologous.
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Meiosis
2. Assemble a second pair of replicated sister chromatids; this time using 12 beads, instead of 20, per pair (six beads per each complete sister chromatid strand).
3. Pair up the homologous chromosome pairs created in Step
1 and 2. DO NOT SIMULATE CROSSING OVER IN THIS
TRIAL. You will simulate crossing over in Part 2.
5. Configure the chromosomes as they would appear in each of the stages of meiotic division (prophase I and II, meta- Figure 4: Second set of replicated chromophase I and II, anaphase I and II, telophase I and II, and somes. cytokinesis). 6. Diagram the corresponding images for each stage in the sections titled “Trial 1 - Meiotic Division
Beads Diagram”. Be sure to indicate the number of chromosomes present in each cell for each phase.
7. Disassemble the beads used in Part 1. You will need to recycle these beads for a second meiosis trial in Steps 8 - 13.
Trial 1 - Meiotic Beads Diagram:
Prophase I
Metaphase I
Anaphase I
Telophase I
Prophase II
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Meiosis
Metaphase II
Anaphase II
Telophase II
Cytokinesis
Part 2: Modeling Meiosis with Crossing Over
8. Build a pair of replicated, homologous chromosomes. 10 beads should be used to create each individual sister chromatid (20 beads per chromosome pair). Two five-holed beads represents the centromere. To do this...
a. Start with 20 beads of the same color to create your first sister chromatid pair. Five beads must be snapped together for each of the four different strands. Two strands create the first chromatid, and two strands create the second chromatid with a 5-holed bead at the center of each chromatid. This creates an “I” shape.
b. Connect the “I” shaped sister chromatids by the 5-holed beads to create an “X” shape.
c. Repeat this process using 20 new beads (of a different color) to create the second sister chromatid pair.
9. Assemble a second pair of replicated sister chromatids; this time using 12 beads, instead of 20, per pair
(six beads per each complete sister chromatid strand). Snap each of the four pieces into two five-holed beads to complete the set up.
10. Pair up the homologous chromosomes created in Step 8 and 9.
11. SIMULATE CROSSING OVER. To do this, bring the two homologous pairs of sister chromatids together
(creating the chiasma) and exchange an equal number of beads between the two. This will result in chromatids of the same original length, there will now be new combinations of chromatid colors.
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Meiosis
12. Configure the chromosomes as they would appear in each of the stages of meiotic division (prophase I and II, metaphase I and II, anaphase I and II, telophase I and II, and cytokinesis).
13. Diagram the corresponding images for each stage in the section titled “Trial 2 - Meiotic Division Beads
Diagram”. Be sure to indicate the number of chromosomes present in each cell for each phase. Also, indicate how the crossing over affected the genetic content in the gametes from Part1 versus Part 2.
Trial 2 - Meiotic Division Beads Diagram:
Prophase I
Metaphase I
Anaphase I
Telophase I
Prophase II
Metaphase II
Anaphase II
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Meiosis
Telophase II
Cytokinesis
Post-Lab Questions
1. In this experiment, how many chromosomes were present when meiosis I started?
2. In this experiment, how many nuclei are present at the end of meiosis II? How many chromosomes are in each?
3. What is the ploidy of the DNA at the end of meiosis I? What about at the end of meiosis II?
4. How are meiosis I and meiosis II different? List two reasons.
5. Why do you use non-sister chromatids to demonstrate crossing over?
6. What combinations of alleles could result from a crossover between BD and bd chromosomes?
7. Identify two ways that meiosis contributes to genetic recombination.
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Meiosis
8. Why is it necessary to reduce the number of chromosomes in gametes, but not in other cells?
9. Blue whales have 44 chromosomes in every cell. Determine how many chromosomes you would expect to find in the following:
Sperm Cell:
Egg Cell:
Daughter Cell from Mitosis:
Daughter Cell from Meiosis II:
10. Research and find a disease that is caused by chromosomal mutations. When does the mutation occur?
What chromosomes are affected? What are the consequences?
11. Diagram what would happen if sexual reproduction took place for four generations using diploid (2n) cells. 191
Lab 13
DNA and RNA
DNA and RNA
Learning Objectives
•
Compare and contrast the structure and function of DNA and RNA
•
Connect nucleotides, amino acids and proteins to the central dogma of the genetic code
•
Explain and perform a DNA extraction
Introduction
Long before scientists had any understanding of how humans acquire their physical traits, they knew that characteristics were passed on from generation to generation. Scientists also knew that traits were expressed as heritable proteins, but they had no idea what mechanism caused these patterns. However, they did know that there were three primary criteria which needed to be met. These criteria were:
1. It needed to carry information between generations.
2. It needed to express that information.
3. It needed to be easily replicable.
DNA Structure
Prior to the 1950’s, there was much debate over what the structure of a molecule that met all three criteria would look like. Though a number of people made significant contributions, in 1953 James Watson and Francis Crick won the Nobel Prize for their model of what we now know as DNA (deoxyribonucleic acid). The features of this model satisfied all of the necessary criteria.
DNA takes the form of what is commonly referred to as a “double helix”, and is often described as looking like a long twisted ladder with rungs
(Figure 1). The sides of the ladder consist of a sugar phosphate
“backbone” and the strand itself has directionality. In other words, like the words on this page, there is a set order in which a genetic information is read. DNA, for example, is read from the 5’ (five prime) to
3’ (three prime) end.
C
G
A
T
G
C
The rungs of the ladder carry information in a sequential series of four
A
different nucleotides (small molecules): Guanine (G), Adenine (A), ThyT mine (T) and Cytosine (C). These nucleotides pair up with each other
Figure 1: The hydrogen bonds formed via hydrogen bonding. A forms two hydrogen bonds with T, and G between adenine and thymine and forms three hydrogen bonds with C (Figure 1). No other combinations cytosine and guanine contribute to are ever made because of the chemical and electrical forces within the the double-helix structure of DNA. nucleotides. 195
DNA and RNA
Genes exists within chromatin, are stretches nucleotides which encode ribonucleic acid (RNA) and protein products. They are often described as units of heredity. Genes have specific locations on the
DNA strand and code for heritable traits (such as hair color). The human genome is comprised of 23 pairs of homologous chromosomes.
Homologous chromosomes carry the same genes but often different versions, or, alleles. Alleles are alternative forms of the same gene (e.g., brown vs. blue eyes). Somatic cells (e.g., bone, heart, skin, liver) contain two alleles (2n) for each gene. One allele is supplied by the parent sperm cell, and the second is supplied by the egg cell. ?
Did You Know...
Although “dogmas” are typically associated with a philosophy or belief system, The Central Dogma refers to the hypothesis that describes biological processes of transcription and translation. Specifically, The Central
Dogma describes:
DNA
(Transcription) mRNA During interphase of the cell cycle division, the DNA double helix
(Translation)
splits into a single helix (Figure 3). Each single helix then serves as a
Protein
template for a new strand of DNA. Neighboring nucleotides then bind to the single strand helix after which a new sugar-phosphate back- This fundamental process of reading the genetic code and developing probone is formed. teins from that code drives life.
Talk About It!
The specificity in which the nucleotides pair means the two new double helices (DNA) are identical to the original. It is the sequence of Based on what you know so far, do you think this process could function these nucleotides that are passed on from one generation to another,
DNA)?
in reverse (e.g., RNA as heritable information.
So the question remains, why is the sequence of these different four nucleotides so important? Simply put, they instruct your cells what proteins to make and how to make them via genetic sequences. This is a very important responsibility because, for example, humans and many other species are composed of proteins.
Serious health concerns or even fatality may occur if developed proteins are incorrect or product at an inappropriate concentration.
RNA Structure
Ribonucleic acid (RNA), also a nucleic acid, varies in structure and function from DNA. As the name states, RNA contains ribose while
DNA is missing the hydroxyl group (“deoxy”) from its ribose. RNA can be single-stranded or double stranded, but in most instances is singlestranded and is shorter than DNA. In RNA, the nucleotide thymine is replaced by uracil which bonds with adenine. There is more than type of RNA involved in protein synthesis: messenger (mRNA), transfer
(tRNA), and ribosomal (rRNA).
Figure 2: RNA structure.
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DNA and RNA
Transcription
For a cell to create proteins it needs to get coding information from DNA housed in the nucleus to ribosomes in the cytoplasm. To do this, a strand of messenger RNA (mRNA) is created by transcription of a gene located in the cell’s DNA. This mRNA then leaves the nucleus through nuclear pores and travels to a ribosome.
Translation
Proteins are simply chains of amino acids (small molecular building blocks) that are linked together. Twenty different amino acids are available to produce all the proteins in the body. Each amino acid is coded for by a three nucleotide sequence (codon) located on mRNA. The sequence of the amino acids determines the size of the protein and how it will fold, both factors that determine its function. Other factors such as charge and hydrophobicity (an aversion to water molecules) play a role in determining how a protein folds. Consider the following analogy:
“The Earth revolves around the sun.”
•
Figure 3: Proteins (a) are chains of amino acids asEach of the 12 different letters (codons) in the preceding sentence is sembled in ribosomes (b).
largely uninformative.
• When letters are assembled they create words, which have meaning.
• Linked words create a sentence (protein), which is then informative.
Consider a protein that is five amino acids long. Picking from the 20 available amino acids there are 520 different possible combinations (3,200,000). Even small proteins are typically several hundred amino acids long.
The number of different proteins that can ultimately be coded for by 20 amino acids is virtually limitless.
Figure 4: Gene expression; the central dogma of molecular biology.
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DNA and RNA
The next obvious question is how do the four nucleotides “code” for 20 different amino acids. Each
“letter” (codon) in the genetic code is made up of three nucleotides which codes for a specific amino acid. If we start with four possible nucleotides (A, T, G, C), how does your body make twenty different amino acids?
• If the “letter” is two nucleotides long, there are 16 possible “letters” (24) - This is not enough options.
• If the “letter” is three nucleotides long, there are 64 possible combinations (34) - more than what’s needed for twenty amino acids.
Like a sentence, the reader (a cell) needs to know where to start and where to stop (two more codons, for a total of 22). The remaining 42 possible combinations make up what is referred to as “the redundancy of the code”. In other words; Tim, Tom, Tam would all be the same person, it is simply three different spellings for his name. Each combination of three nucleotides is known as a “codon”.
Second Letter
U
C
UUU
U
UUC
UCU
Phe
UUA
UUG
Leu
First Letter
Leu
UCG
CCU
CCC
CCA
CUG
CCG
AUU
Ile
AUA
AUG
Met
GUC
UGC
Cys
C
Stop
UGA
Stop
A
UAG
Stop
UGG
Trp
G
CAU
His
CGU
CAC
Pro
CAA
CGC
Gln
U
Arg
C
CGA
A
CAG
CGG
G
ACU
AAU
AGU
U
ACC
AAC
ACA
GUU
G
UAA
Ser
U
UGU
UAC
UCC
CUA
AUC
UAU
Thr
AAA
ACG
AAG
GCU
GAU
GCC
Val
Asn
AGC
Ser
AGA
Lys
Asp
GAC
AGG
A
Arg
GGU
G
U
GGC
Ala
C
C
Gly
GUA
GCA
GAA
GUG
GCG
GAG
Glu
GGA
A
GGG
G
Figure 5: Chart showing how codons code for specific amino acids.
198
Third Letter
A
CUC
G
Tyr
UCA
CUU
C
A
DNA and RNA
Pre-Lab Questions
1. Arrange the following molecules from least to most specific with respect to the original nucleotide sequence: RNA, DNA, Amino Acid, Protein
2. Identify two structural differences between DNA and RNA.
3. Suppose you are performing an experiment in which you must use heat to denature a double helix and create two single stranded pieces. Based on what you know about nucleotide bonding, do you think the nucleotides will all denature at the same time? Use scientific reasoning to explain why.
Experiment 1: Coding
In this experiment, you will model the effects of mutations on the genetic code. Some mutations cause no structural or functional change to proteins while others can have devastating affects on an organism.
Materials
Red Beads
Yellow Beads
Blue Beads
Green Beads
Procedure
1. Using the red, blue, yellow and green beads, devise and lay out a three color code for each of the following letters (codon). For example Z = green : red : green.
In the spaces below the letter, record your “code”.
C:
___
E:
___
H:
___
I:
K:
L:
___
___
___
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DNA and RNA
M:
___
Create codons for:
O:
S:
T:
U:
___
___
___
___
Start:
Stop:
Space:
___
___
___
2. Using this code, align the beads corresponding to the appropriate letter to write the following sentence
(don’t forget start, space and stop): The mouse likes most cheese
a. How many beads did you use?
There are multiple ways your cells can read a sequence of DNA and build slightly different proteins from the same strand. We will not go through the process here, but as an illustration of this “alternate splicing”, remove codons (beads) 52 - 66 from your sentence above.
b. What does the sentence say now? (re-write the entire sentence)
Mutations are simply changes in the sequence of nucleotides. There are three ways this occurs:
1. Change a nucleotide(s)
2. Remove a nucleotide(s)
3. Add a nucleotide(s)
3. Using the sentence from exercise 1B:
a. Change the 24th bead to a different color. What does the sentence say now (re-read the entire sentence)? Does the sentence still make sense?
b. Replace the 24th bead and remove the 20th bead (remember what was there). What does the sentence say (re-read the entire sentence)? Does the sentence still make sense? If it doesn’t make sense as a sentence, are there any words that do? If so, what words still make sense?
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DNA and RNA
c. Replace the 20th bead and add one between bead numbers 50 and 51. What does the sentence say now? Does the sentence still make sense?
d. In 3.a (above) you mutated one letter. What role do you think the redundancy of the genetic code plays in this type of change?
e. Based on your observations, why do you suppose the mutations we made in 3.b and 3.c are called frame shift mutations?
f.
Which mutations do you suspect have the greatest consequence? Why?
Experiment 2: Transcription and Translation
DNA codes for all of the proteins manufactured by any organism (including you!). It is valuable, highly informative and securely protected in the nucleus of every cell. Consider the following analogy:
An architect spends months or years designing a building. Her original drawings are valuable and informative. She will not provide the original copy to everyone involved in constructing the building. Instead, she gives the electrician a copy with the information she needs to build the electrical system. She will do the same for the plumbers, the framers, the roofers and everyone else who needs to play a role to build the structure. These are subsets of the information contained in the original copy. Your cell does the same thing. The “original drawings” are contained in your DNA which is securely stored in the nucleus.
Nuclear DNA is “opened up” by an enzyme called helicase, and a subset of information is transcribed into
RNA. RNA is a single strand version of DNA, where the nucleotide uracil, replaces thymine. The copies are sent from the nucleus to the cytoplasm in the form of messenger RNA (mRNA ). Once in the cytoplasm, transfer RNA (tRNA) links to the codons and aligns the proper amino acids, based on the mRNA sequence.
Protein builders called ribosomes float around in the cytoplasm, latch onto the strand of mRNA and sequentially link the amino acids together that the tRNA has lined up for them. This construction of proteins from the mRNA is known as translation.
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DNA and RNA
Materials
Blue beads
*You Must Provide
Green beads
Red beads
In this experiment:
Yellow beads
•
Regular beads are used as nucleotides.
Pop-it® beads (8 different colors)
•
Pop-it® beads are used as amino acids.
*Pen or pencil
Procedure
1. Use a pen or pencil to write a five word sentence using no more than eight different letters in the space below. 2. Now, use the red, blue, green, and yellow beads to form “codons” (three beads) for each letter in your sentence. Then, create codons to represent the “start, “space” and stop” regions within your sentence.
Write the sentence using the beads in the space below:
3. How many beads did you use?
4. Assign one Pop-It® bead to represent each codon. You do not need to assign a Pop-It® bead for the start, stop and space regions. These will be your amino acids.
5. Connect the Pop-It® beads to build the chain of amino acids that code for your sentence (leave out the start, stop, and space regions).
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DNA and RNA
6. How many different amino acids did you use?
7. How many total amino acids did you use?
Experiment 3: DNA Extraction
Much can be learned from studying an organism’s DNA. The first step to doing this is extracting DNA from cells. In this experiment, you will isolate DNA from the cells of fruit.
Materials
(1) 10 mL Graduated Cylinder
nana, etc.)
(2) 100 mL Beakers
*Scissors
15 cm Cheesecloth
**DNA Extraction Solution
1 Resealable Bag
***Ice Cold Ethanol
1 Rubber Band (Large. Contains latex; please wear gloves when handling if you have a latex allergy).
*You Must Provide
Standing Test Tube
**Contains sodium chloride, detergent and water
Wooden Stir Stick
***For ice cold ethanol, store in the freezer 60 minutes before use.
*Fresh, Soft Fruit (e.g., Grapes, Strawberries, Ba-
Procedure
1. If you have not done so, prepare the ethanol by placing it in a freezer for approximately 60 minutes.
2. Put pieces of a soft fruit into a plastic zipper bag and mash with your fist. The amount of food should be equal to the size of approximately five grapes.
3. Use the 10 mL graduated cylinder to measure 10 mL of the DNA Extraction Solution. Transfer the solution from the cylinder to the bag with the fruit it in. Seal the bag completely.
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DNA and RNA
4. Mix well by kneading the bag for two minutes.
5. Create a filter by placing the center of the cheesecloth over the mouth of the standing test tube, pushing it into the tube about two inches, and securing the cheesecloth with a rubber band around the top of the test tube.
6. Cut a hole in the corner of the bag and filter your extraction by pouring it into the cheesecloth. You will need to keep the filtered solution which passes through the cheese cloth into the standing test tube.
7. Rinse the 10 mL graduated cylinder, and measure five mL of ice-cold ethanol. Then, while holding the standing test tube at a 45° angle, slowly transfer the ethanol into the standing test tube with the filtered solution. 8. DNA will precipitate (come out of solution) after the ethanol has been added to the solution. Let the test tube sit undisturbed for 2 - 5 minutes. You should begin to see air bubbles form at the boundary line between the ethanol and the filtered fruit solution. Bubbles will form near the top, and you will eventually see the DNA float to the top of the ethanol.
9. Gently insert the stir stick into the test tube. Slowly raise and lower the tip several times to spool and collect the DNA. If there is an insufficient amount of DNA available, it may not float to the top of the solution in a form that can be easily spooled or re-
Figure 6: DNA extraction. The color has
moved from the tube. However, the DNA will still be visible as been enhanced by dying the fruit with a white/clear clusters by gently stirring the solution and pushing substance that glows under black light. the clusters around the top.
Post-Lab Questions
1. What is the texture and consistency of the DNA?
2. Why did we use a salt in the extraction solution?
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DNA and RNA
3. Is the DNA soluble in the aqueous solution or alcohol?
4. What else might be in the ethanol/aqueous interface? How could you eliminate this?
5. Which DNA bases pair with each other? How many hydrogen bonds are shared by each pair?
6. How is information to make proteins passed on through generations?
7. Watch the following Virtual Lab demonstrating DNA Extraction. In this experiment, how do the Lysis Solution and the Salt Solution vary by function?
8. Identify one step which was included in the Virtual Lab which was not required in the hands-on experiment. Then, identify one step which was included in the hands-on experiment, but not the virtual lab. Why weren’t these steps required?
205
Lab 14
Mendelian Genetics
Mendelian Genetics
Learning Objectives
•
Explain how Mendel’s work formed the foundation of modern genetics including the law of segregation and the law of independent assortment
•
Relate genes to homozygous, heterozygous, dominant alleles, recessive alleles, genotype and phenotype •
Use monohybrid and dihybrid crosses to analyze patterns of inheritance including dominance, incomplete dominance and co-dominance
Introduction
In 1866, Gregor Mendel (an Austrian Monk), published a paper entitled “Experiments in Plant Hybridization”.
It went largely unnoticed until 1900 when it was “rediscovered” and subsequently became the basis for what we now refer to as Mendelian Genetics. Mendel was the first to recognize:
•
Inherited characteristics are determined by specific factors (now recognized as genes).
•
These factors occur in pairs (homologous genes).
•
When gametes form, these factors segregate so that each gamete contains only one.
The Law of Independent Assortment
These original observations lead to what we now refer to as
The law of segregation and the law of independent assortment. The law of segregation states that during meiosis, homologous (paired) chromosomes split. The law of independent assortment states that during meiosis, each homologous chromosome has an equal chance of ending up in either gamete, and alleles for individual genes segregate with the chromosomes on which they are located (Figure 1).The law of independent assortment allows for genetic recombination. The following equation can be used to determine the total number of Figure 1: Law of Independent Assortment. possible genotype combinations for any particular number of genes: 2g= Number of possible genotype combinations (where “g” is the number of genes)
•
1 gene: 21= 2 genotypes
•
2 genes:22= 4 genotypes
•
3 genes: 23 = 8 genotypes
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Genotype and Phenotype
Genotype refers to the combination of alleles for a particular trait. Phenotype refers to the appearance of that combination of alleles. In other words, genotype determines phenotype. Genotype cannot be seen and is represented by letters. Using corn as an example (Figure 2):
•
The large chromosome has the gene for kernel color (Y = yellow, y = blue).
•
The small chromosome has the gene for kernel texture (S = smooth (green); s = wrinkled (red)).
Dominance
In our example, the genotype of the diploid cell is Yy, Ss, while the phenotype is Yellow and Smooth. When a dominant allele is present, that characteristic is expressed, regardless of the second allele. In this case both the Yy and
YY offspring will be yellow. The phenotype inscribed by a recessive allele is only expressed when both alleles are recessive. In this case, only the yy combination is blue. The dominant allele is always represented by capital letters, while the recessive is represented by lower case letters. Any letter Figure 2: If one dominant allele is present the can be used to represent a gene, but usually the first letter of dominant phenotype is expressed. If two recessive alleles combine a recessive phenotype is the dominant phenotype used (ex., Yellow). expressed. Homozygous and Heterozygous Genotypes
When both alleles of a gene are the same they are said to be homozygous. If they are different they are said to be heterozygous. When gametes form, these factors segregate so that each gamete contains only one allele for each gene. Remember, alleles are one copy of a gene and are coded within a chromosome.
Incomplete Dominance and Co-Dominance
Alleles can exhibit incomplete dominance and co-dominance. An example of incomplete dominance is the cross of two plants, one with red flowers and one with white, whose offspring have pink flowers. In the case of co-dominance, the same cross would result in red and white striped flowers.
Predicting Inheritance
If we know the genotype of two parents we can predict both the genotype and phenotype of their offspring using a Punnett Square. A monohybrid cross is a cross between two parents (P), looking at a single gene. In this example, both parents are pure breeding (homozygous); one for the yellow color and one for the blue color. This cross can be shown as a Punnett Square (Figure 3), with each parent (P) contributing a single gamete. The offspring (F1) are determined by adding the gamete of each parent (P) (Row and Column). The cross of the (F1) generation, known as the F2 generation, is shown in Figure 4.
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When the P generation is crossed one hundred percent offspring are heterozygous (Y , y) and yellow in appearance even though one parent had blue kernels (Figure 3). When two offspring from the F1 generation are crossed their offspring have a 75% chance of having yellow kernels and 25% chance of having blue kernels (Figure 4). While the recessive phenotype disappears in the F1 generation it reappears in the F2 generation. Parent 1
Y
Y
y
Y y
Y y
y
Y y
Y y
Parent
2
Figure 3: Punnett Square Monohybrid Cross F1.
The Punnett square can also predict the F1 for multiple genes. Using our corn example, let’s look at two genes (color and texture), also known as a dihybrid cross. In this example we use two parents that are heterozygous for both traits (Figure 5), using the gametes we already identified in (Figure 3).
Parent 1
Y
y
Y
Y Y
Y y
y
Y y
y y
Parent
2
Figure 4: Punnett Square Monohybrid Cross F2.
The F2 phenotypes are:
Yellow & Smooth: 9
Yellow & Wrinkled: 3
Blue & Smooth: 3
Blue & Wrinkled: 1
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Mendelian Genetics
Y s
Y S
y S
y s
Y s
Y Y s s
Y Y S s
y Y S s
y Y s s
Y S
Y Y s S
Y Y S S
y Y S S
y Y s S
y S
Y y s S
Y y SS
y y S S
y y s S
y s
Y y s s
Y y S s
y y S s
y y s s
Figure 5: Punnett square of a dihybrid cross (F1).
Pre-Lab Questions
1. In a species of mice, brown fur color is dominant to white fur color. When a brown mouse is crossed with a white mouse all of their offspring have brown fur. Why did none of the offspring have white fur?
2. Can a person’s genotype be determine by their phenotype? Why or why not?
3. Are incomplete dominant and co-dominant patterns of inheritance found in human traits? If yes, give examples of each.
4. Consider the following genotype: Yy Ss Hh. We have now added the gene for height: Tall (H) or Short (h).
a. How many different gamete combinations can be produced?
b. Many traits (phenotypes), like eye color, are controlled by multiple genes. If eye color were controlled by the number of genes indicated below, how many possible genotype combinations would there be in the following scenarios?
5 Eye Color Genes:
10 Eye Color Genes:
20 Eye Color Genes:
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Mendelian Genetics
Experiment 1: Punnett Square Crosses
In this experiment you will use monohybrid and dihybrid crosses to predict patterns of inheritance.
Materials
Blue Beads
Yellow Beads
Green Beads
(2) 100 mL Beakers
Red Beads
Permanent Marker
Procedure
Part 1: Punnett Squares
1. Set up and complete Punnett squares for each of the following crosses: (remember Y = yellow, and y = blue) Y Y and Y y
Y Y and y y
2. What are the resulting phenotypes?
3. Are there any blue kernels? How can you tell?
4. Set up and complete a Punnett squares for a cross of two of the F1 from Step 1 (above).
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5. What are the genotypes of the F2 generation?
6. What are their phenotypes?
7. Are there more or less blue kernels than in the F1 generation?
8. Identify the four possible gametes produced by the following individuals:
a) YY Ss:
______
______
______
______
b) Yy Ss:
______
______
______
______
c) Create a Punnett square using these gametes as P and determine the genotypes of the F1:
What are the phenotypes? What is the ratio of those phenotypes?
Part 2 and 3 Setup
1. Use the permanent marker to label the two 100 mL beakers as “1” and “2”.
2. Pour 50 of the blue beads and 50 of the yellow beads into Beaker 1. Sift or stir the beads around to create a homogenous mixture.
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3. Pour 50 of the red beads and 50 of the green beads into Beaker 2. Sift or stir the beads around to create a homogenous mixture.
Assumptions for the remainder of the experiment:
•
Beaker 1 contains beads that are either yellow or blue.
•
Beaker 2 contains beads that are either green or red.
•
Both beakers contain approximately the same number of each colored bead.
•
These colors correspond to the following traits (remember that Y/y is for kernel color and S/s is for smooth/wrinkled):
1. Yellow (Y) vs. Blue (y)
2. Green (G) vs. Red (g).
Part 2: Monohybrid Cross
1. Randomly (without looking) take two beads out of Beaker 1. This is the genotype of Individual #1. Record the genotype in Table 1. Do not put these beads back into the beaker.
Table 1: Parent Genotypes: Monohybrid Crosses
Genotype of In-
Genotype of Indi-
P1
P2
P3
P4
Generation
P
2. Repeat Step 1 for Individual #2. These two genotypes represent the parents (generation P) for the next generation. 3. Set up a Punnett square and determine the genotypes and phenotypes for this cross. Record your data in Table 2
4. Repeat Step 3 four more times (for a total of five subsequent generations). Return the beads to their respective beakers when finished.
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Table 2: Generation Data Produced by Monohybrid Crosses
Parents
Possible Offspring Genotypes
Possible Offspring Phenotypes
Genotype
Ratio
Phenotype
Ratio
P
P1
P2
P3
P4
Part 3: Dihybrid Cross
1. Randomly (without looking) remove two beads from of Beaker 1 and two beads from Beaker 2. These four beads represent the genotype of Individual #1. Record this information in Table 3
2. Repeat Step 1 to obtain the genotype of Individual #2. Record the phenotypes of both individuals in Table 3.
Table 3: Parent Genotypes: Dihybrid Crosses
Genotype of In-
Genotype of Indi-
P1
P2
P3
P4
Generation
P
3. Determine what the possible genotypes might be if each individual produced gametes. Record these possible genotypes in Table 4.
4. Predict the ratio of possible genotypes which could be produced by a cross between Individual #1 and
Individual #2? Record your predictions in Table 4.
Hint: Think back to the example the dihybrid cross in the Introduction. Record your prediction in Table
2.
5. Set up a Punnett square and determine the genotypes and phenotypes for this cross.
6. Repeat Step 5 four additional times (for a total of five subsequent generations).
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Table 4: Generation Data Produced by Dihybrid Crosses
Parents
Possible Offspring Genotypes
Possible Offspring Phenotypes
Genotype
Ratio
Phenotype
Ratio
P
P1
P2
P3
P4
Post-Lab Questions
Part 2: Monohybrid Cross
1. How much genotypic variation do you find in the randomly picked parents of your crosses?
2. How much in the offspring?
3. How much phenotypic variation?
4. Is the ratio of observed phenotypes the same as the ratio of predicted phenotypes? Why or why not?
5. Pool all of the offspring from your five replicates. How much phenotypic variation do you find?
6. What is the difference between genes and alleles?
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Mendelian Genetics
7. How might protein synthesis execute differently if a mutation occurs?
8. Organisms heterozygous for a recessive trait are often called carriers of that trait. What does that mean?
9. In peas, green pods (G) are dominant over yellow pods. If a homozygous dominant plant is crossed with a homozygous recessive plant, what will be the phenotype of the F1 generation? If two plants from the F1 generation are crossed, what will the phenotype of their offspring be?
Part 3: Dihybrid Cross
1. How similar are the observed phenotypes in each replicate?
2. How similar are they if you pool your data from each of the five replicates?
3. Is it closer or further from your prediction?
4. Did the results from the monohybrid or dihybrid cross most closely match your predicted ratio of phenotypes?
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Mendelian Genetics
5. Based on these results; what would you expect if you were looking at a cross of 5, 10, 20 independently sorted genes?
6. Why is it so expensive to produce a hybrid plant seed?
7. In certain bacteria, an oval shape (O) is dominant over round (o) and thick cell walls (T) are dominant over thin (t). Show a cross between a heterozygous oval, thick cell walled bacteria with a round, thin cell walled bacteria. What are the phenotype of the F1 and F2 offspring?
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Lab 15
Population Genetics
Population Genetics
Learning Objectives
•
Explain how genetic drift, the founder effect, mutations and natural selection affect a population’s gene pool
•
Use the Hardy-Weinberg equation to calculate gene frequencies within a population
•
Analyze the effects of stochastic events on genetic variation and frequency in a gene pool
Introduction
In the previous lab we looked at how genes are passed on to offspring. In this lab, the exercises are designed to look at individual genes (two alleles, one dominant, one recessive). However, we will be looking at their presence, prevalence and distribution at the population level.
Gene Frequency
The gene pool is the sum of all genes and their corresponding alleles in a given population. Take a look at the population of 100 brown and white mice in Figure 1. The color brown (B) is dominant. The standard for naming alleles is to use the case of the dominant trait, with the lower case to represent the recessive allele.
Their gene pool is B, b.
Gene frequency refers to how many times each allele is found in the population. These 100 mice have 200 alleles: •
55 heterozygous mice (B, b) have 55 B alleles and 55 b alleles.
•
27 homozygous recessive mice (b, b) have 54 b alleles (2 x 27 “b”= 54).
•
18 homozygous dominant mice (B, B) have 36 B alleles (18 x 2 “B” = 36).
The gene frequency of the population is:
B: 91 b: 109
Figure 1: Mouse population.
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Population Genetics
Often this is represented as a percentage of the dominant gene. In this case, the percentage of B is 45.5%
(=91/200). Note that the dominant gene is less prevalent than the recessive gene. This is not unusual. It is important to remember that dominance has no direct relationship to prevalence.
Hardy-Weinberg Equilibrium
To estimate the frequency of alleles in a population, we can use the Hardy-Weinberg equation. According to this equation: p = the frequency of the dominant allele (represented here by A) q = the frequency of the recessive allele (represented here by a)
For a population in genetic equilibrium:
p + q = 1.0 (The sum of the frequencies of both alleles is 100%.)
(p + q)2 = 1
So
p2 + 2pq + q2 = 1
The three terms of this binomial expansion indicate the frequencies of the three genotypes:
p2 = frequency of AA (homozygous dominant)
2pq = frequency of Aa (heterozygous) q2 = frequency of aa (homozygous recessive)
Stochastic Events
Many stochastic events change both the gene pool and gene frequencies over time. These parameters can also change as a result of mutation and natural selection.
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Population Genetics
Mutations are a change in the sequence of DNA. Most mutations do not change the phenotype and confer no advantages or disadvantages to the individual. Each of us has hundreds and probably thousands of mutations.
Nonetheless, some mutations do change the protein coded for by a gene. The vast majority of these mutations are lethal and the embryo never fully develops. Occasionally mutations do not effect embryonic development and the offspring is born without complication.
Pre-Lab Questions
Assumptions:
•
There are approximately 3,000,000,000 base pairs in the mammalian genome (genes constitute only a portion of this total).
•
There are approximately 10,000 genes in the mammalian genome.
•
A single gene averages 10,000 base pairs in size.
•
Only 1 out of 3 mutations that occur in a gene result in a change to the protein structure.
In the mammalian genome:
1. How many total base-pairs are in all the mammalian genes?
2. What proportion (%) of the total genome does this represent?
3. What is the probability that a random mutation will occur in any given gene?
4. What is the probability that a random mutation will change the structure of a protein
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Population Genetics
Note: In the following experiments on gene pool, gene frequency, and genetic diversity; assume there are four alleles for color and that they are all homologous.
Experiment 1: Genetic Variation
Genetic variation is simply the genetic difference within or between populations, in the gene pool and/or gene frequency.
Consider the following two populations of butterflies (Figure 2):
Figure 2: Butterfly populations.
Assumptions: Both populations contain the same four colors of butterflies, thus the gene pool is the same.
However, the distribution of colors within that population is different, thus their gene frequencies are different.
Materials
Blue Beads
Yellow Beads
Red Beads
(2) 100 mL Beakers
Green Beads
(2) 250 mL Beakers
Procedure
1. Pour 50 blue beads and 50 red beads into a 250 ml beaker. Without looking, randomly take 50 beads from the 250 mL beaker and place them in a 100 mL beaker (this is Beaker #1).
2. Pour 50 green beads and 50 yellow beads into a second 250 mL beaker. Without looking, randomly take
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Population Genetics
20 beads from the 250 mL beaker and place them in the other 100 mL beaker (this is beaker #2).
Note: When done, return beads to their respective beakers (1 or 2) for use in the next experiment.
Post-Lab Questions
1. What is the gene pool of beaker #1?
2. What is the gene pool of beaker #2?
3.
What is the gene frequency of beaker #1?
4.
What is the gene frequency of beaker #2?
5. What can you say about the genetic variation between these populations?
Experiment 2: Genetic Drift
Genetic drift, the variation of the gene pool and/or gene frequency of a population, can result from a variety of stochastic (random) means. Consider the following population of butterflies who have half of their habitat destroyed by wildfire (Figure 3):
Figure 3: Butterfly populations before and after wildfire.
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Population Genetics
The remaining population has 50% of the initial gene pool (two colors) and the gene frequency is different. As these individuals reproduce, their offspring will no longer reflect the original population.
Materials
Blue Beads
Beads in Beaker #1 (from Experiment 1)
Green Beads
Beads in Beaker #2 (from Experiment 1)
Red Beads
(1) 100 mL Beaker
Yellow Beads
(1) 250 mL Beaker
Procedure
1. From the 250 mL beaker containing green and yellow beads, take 10 beads and place them into the unused 100 mL beaker (this is Beaker #3).
2. Randomly remove half of the beads from Beakers #1, #2, and #3. These are the individuals that survived the fire. Keep them separated so they can be returned to their proper beaker. You will use them again in the next lab.
3. Record your results and place the beads back in their respective beakers.
4. Repeat this process four more times (five total).
Post-Lab Questions
1. What observations can you make regarding the gene pool and gene frequency of the surviving individuals?
2. Do the results vary between the populations represented by beakers #1, #2 and #3? Why or why not?
3. What observations can you make about the genetic variation between the parent and surviving populations?
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Population Genetics
Experiment 3: Stochastic Events
Consider the same population of butterflies is in the path of a hurricane. All survive, but 10 are blown to a new location. These 10 start a new population; their progeny will reflect the founder’s gene pool. This is known as the founder effect.
Figure 4: Butterfly populations before and after hurricane.
Materials
Beaker #1 (from Experiment 1)
Beaker #2 (from Experiment 1)
Beaker #3 (from Experiment 2)
Procedure
1. Remove 10 individuals from Beaker #1, five from Beaker #2 and two from Beaker #3. These are the founders of your new population.
2. Record your results and place the beads back in their respective beakers.
3. Repeat this process four more times (five total).
Note: When you are finished with this experiment, return the beads to their appropriate bags.
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Post-Lab Questions
1. What observations can you make regarding the gene pool and gene frequency of the founding individuals?
2. Do these results vary between the populations founded by beakers #1, #2 and #3? Why or why not?
3. What observations can you make about the genetic variation between the parent and founding populations?
4. Suppose you have a population of 300 butterflies. If the population experiences a net growth of 12% in the following year, how many butterflies do you have?
5. Now suppose you have 300 eggs, but only 70% of those eggs progress to become a caterpillar, and only 80% of the caterpillar progress to become an adult butterfly. How many butterflies do you have?
(For simplicity, assume that all butterflies survive to the next year, in this example)
6. Suppose you have a population of 150 butterflies, but a wildfire devastates the population and only 24 butterflies survive. What percent does the colony decrease by?
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Population Genetics
Experiment 4: Natural Selection
Natural selection is a selection pressure that effects phenotypes in one of three ways:
1. It will create an adaptive advantage.
2. It will create an adaptive disadvantage.
3. It will remain entirely neutral.
A classic example to illustrate natural selection comes from England. Prior to the Industrial Revolution, the native moths were normally a light color, though darker versions of the same species existed. The lighter color blended with the light bark of the local trees, while the darker moths experienced a higher predation rate - they were easier for birds to spot and fewer survived to reproduce. As England entered the Industrial
Revolution they began burning fossil fuels with little regard to the pollutants they were emitting. The trunks of the trees became coated with soot and their color darkened. The lighter moths became more conspicuous and the darker moths were better camouflaged. The proportion of white to dark moths changed.
Materials
(1) 100 mL Beaker
Red Beads
Blue Beads
Access to a Printer
Procedure
1. Print the two sheets of paper marked Blue Habitat (Figure 5) and Red Habitat (Figure 6), from your manual (found at the end of this procedure).
2. Place 50 red and 50 blue beads into a 100 mL beaker.
3. Mix them well and pour them onto the sheet marked Red Habitat.
4. Keep the beads that fall onto habitat that matches their color.
5. For each bead that you keep (and return to the beaker), add another bead of the same color to the beaker (discard the rest).
6. Repeat this three times.
7. Record the remaining colors.
Blue _____________ Red _____________
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Population Genetics
Do you observe a selective advantage for the red or blue beads? Why?
8. Repeat the process using the Blue Habitat with the remaining beads.
What beads remain now?
Blue _____________
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Red _____________
Population Genetics
Figure 5: Red Habitat
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Figure 6: Blue Habitat
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Post-Lab Questions
1. How did the distribution of phenotypes change over time?
2. Is there a selective advantage or disadvantage for the red and/or blue phenotypes?
3. What phenotypic results would you predict if starting with the following population sizes?
A. 1000:
B. 100:
C. 10:
4. Assume that you live in a country with 85 million people that consistently experiences an annual growth rate of 4.2%. If this population continues to grow at the same rate for the next 5 years, how many people will live in the country (round to the nearest whole number).
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Population Genetics
Experiment 5: Sickle Cell Anemia Inheritance Patterns
Sickle cell anemia is a genetic disease (one base pair mutation that changes a protein). It is more common in those of African ancestry. “S” will represent the normal dominant allele and “s” the recessive sickle allele.
They are co-dominant alleles – SS is normal, Ss is not fatal, ss is debilitating, painful and often fatal.
Materials
(1) 100 mL Beaker
Blue Beads
Red Beads
Procedure
1. Place 25 red (S) and 25 blue (s) beads into the 100 mL beaker and mix well.
2. Randomly (without looking) remove two beads. Repeat 10 times (without returning the beads to the beaker), each time recording if it was a SS, Ss or ss.
3. Assume that each individual with the ss genotype passed away, and remove ss instances from the population.
4. The remaining beads survived and reproduced.
5. Count how many red and blue beads remained (separately) and place twice that number back in the beaker. 6. Repeat the process seven times.
Post-Lab Questions
1. What is the remaining ratio of alleles?
2. Have any been selected against?
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3. Given enough generations, would you expect one of these alleles to completely disappear from the population? Why or why not?
4. Would this be different if you started with a larger population? Smaller?
5. After hundreds or even thousands of generations both alleles are still common in those of African ancestry. How would you explain this?
6. The worldwide distribution of sickle gene matches very closely to the worldwide distribution of malaria
(Figure 7). What is the significance of this?
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Figure 7: Distribution of Malaria (top) and Sickle Cell trait (bottom).
Lab 16
Taxonomy
Taxonomy
Learning Objectives
•
Apply Linnaean’s classification system to analyze relatedness
•
Explain how advances in taxonomy such as phylogenetics have changed the way organisms are classified •
Compare and contrast binomial nomenclature and common name
Introduction
Taxonomy is the classification (organization) of organisms into related groups. Very early on, scientists recognized the importance of classifying or ranking organisms. In the 1700s, Carl Linnaeus (also known as Carl von Linné or Carolus Linnaeus) developed what is now known as the Linnaean Classification System. At the highest level in this system, everything in nature was divided into three kingdoms: mineral, vegetable, and animal. Linnaeus then ranked sequentially smaller groups within each kingdom as class, order, genus, species and variety. This contribution to science earned him the title “The Father of Taxonomy”. His system is still, in an evolved form, used today.
Starting with domain, there is a continual increase in specificity as organisms are classified into smaller and smaller categories (Figure 1 ). In other words, the categories get smaller in terms of the number of organisms that are included. As illustrated in Figure 1, the Linnaean system classifies organisms into sequential groups:
•
Domain
•
Kingdom
•
Phylum
•
Class
•
Order
•
Family
•
Genus
•
Species
Interestingly, there is not a consensus definition of
‘species’ among scientists at this time. The biological species concept is the most commonly used definition. Basically it defines a species as a group of individuals that can interbreed and create viable offspring. Sub-species are used in some classifications and is generally accepted, but not always included in the Linnaean system.
Figure 1: Taxonomy Tree. To use this tree, one would begin at the bottom and work up.
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Taxonomy
Table 1: Sample Classifications of Humans and a Red Maple Tree
Example
Human Being
Red Maple
Domain
Eukarya
Eukarya
Kingdom
Animalia
Plantae
Phylum
Chordata
Tracheophyta
Class
Mammalia
Angiospermae
Order
Primates
Sapindales
Family
Hominidae
Acerceae
Genus
Homo
Ace
Species
Sapien
Rubrum
A useful tool to remembering the order of the Linnaean classification system is developed by creating a mnemonic phrase using the first letter of each classification. For example, Daring Kids Pick Cauliflower
Over Fresh Grown Strawberries. Can you create any mnemonics to help remember the Linnaean classification system?
Binomial Nomenclature
Carl Linnaeus also helped to standardize the naming of species by introducing the binomial (two name) species name. Binomial nomenclature uses the genus and species as the formal name of all organisms because they are the two most specific levels therefore unique to the organism. For example, human beings are Homo sapiens. Note that the genus is capitalized while species is not and, since these are Latin names, they are italicized. Some organisms can have more than one common name (i.e. cougar, puma and mountain lion). Using binomial nomenclature specifies a species.
Advances in Taxonomy
Taxonomy is in a constant state of flux. Traditional groupings of organisms were dependent on mostly morphological traits, physiological similarities and, to some extent, embryological events. Consider what techniques were available in the 1700s when Carl Linnaeus described his system for ranking life. Gross anatomy could be studied and identified, but most forms of microscopy were not available.
Phylogenetics
With advances in molecular techniques, nucleic acid (DNA) sequences are used to classify organisms based on their evolutionary history. These advancements have given birth to the field of phylogenetics.
Phylogenetics is the field of study that determines the evolutionary relationships among groups of organisms using morphological traits, behavioral information and molecular data. Ribosomal RNA (rRNA) is cur-
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Taxonomy rently one of the genes of choice for phylogenetic studies. rRNA is necessary for the production of proteins and, because all life forms require proteins, it follows that all organisms include rRNA genes. Sequences from specific regions of an rRNA gene demonstrate similarities amongst different organisms, enabling evolutionary relationships to be explored. It is well accepted that the more similar the sequences are between two organisms, the more closely related the organisms. As DNA sequencing techniques improve (and costs decrease) other regions of the genome are also being sequenced and compared.
Hot Topics in Taxonomy
Keep in mind that new techniques aid in classifications and, at the same time, cause previously well accepted classifications to become obsolete. Currently, many sources list three domains: Bacteria, Archaea, and Eukarya. Within Eukarya are four kingdoms: Protista, Plantae, Fungi, and Animalia. The Kingdom Protista has historically been a ‘catch-all’ category for all single-celled eukaryotic organisms; but, not plants, fungi or animals. However, molecular data has revealed distinct relationships within the eukaryotes that has led to suggestions for new kingdoms and “supergroups”. In this proposal, organisms that share a common ancestor are grouped together and organisms that were previously placed in the Kingdom Protista are distributed throughout the new categories. Recall that the organization of organisms based on shared common ancestors is considered phylogeny. The recently suggested revised “supergroups” are Bacteria, Archaea, Excavata, Amoebozoa, Opisthokonta, Rhizaria, Chromalveolata, and Archaeplastida. In this organizational system, animals fall within Opisthokonta and plants are placed in Archaeplastida.
Classifications will continue to evolve as scientists are able to delve at increasingly deeper molecular levels. For now, the Kingdom Protista will continue to be used in this manual as we briefly survey the three domains (Bacteria, Archaea and Eukarya) and four kingdoms (Protista, Plantae, Fungi, and Animalia).
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Taxonomy
Pre-Lab Questions
1. Use the following classifications to determine which organism is least related out of the three. Explain your rationale.
Table 2: Classifications
Classification Level
American Green Tree
Frog
European FireBellied Toad
Eastern Newt
Domain
Eukarya
Eukarya
Eukarya
Kingdom
Animalia
Animalia
Animalia
Phylum
Chordata
Chordata
Chordata
Class
Amphibia
Amphibia
Amphibia
Order
Anura
Anura
Caudata
Family
Hylidae
Bombinatoridae
Salamandridae
Genus
Tursipops
Bombina
Notophthalmus
Species
cinerea
bombina
viridescens
2. How has DNA sequencing affected the science of classifying organisms?
3. You are on vacation and see an organism that you do not recognize. Discuss what possible steps you can take to classify it.
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Taxonomy
Experiment 1: Dichotomous Key Practice
In this experiment, you will identify organisms by their binomial nomenclature using a dichotomous key. A dichotomous key is an identification tool that starts with a broad defining characteristics and splits into two options until an organism can be identified.
Materials
Images of Organisms (Figure 2)
Dichotomous Key (Figure 3)
Procedure
1. Start by observing organism i (Figure 2). Once you have taken notice of its physical characteristics, use the dichotomous key (Figure 3) to identify the organism.
2. Start at number 1 on the key and decide if the organism has feature “1a” or feature “1b”.
3. Which ever option you choose, follow the dotted line over to the next step that will state “Go to #”.
4. Go to that number and again decide between the two options.
5. Eventually you will wind up at a two name scientific name.
6. Once you have identified organism i record your finding in Table 3.
7. Repeat steps 1-6 for all of the organisms in the table.
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Taxonomy
i.
ii.
iii.
iv.
vi.
v.
vii.
viii. ix. x.
xi.
xiii. xii. 246
Figure 2: Organisms to be identified.
Taxonomy
1a. Animal…………………………………………………………………………………..………………………………………………………………………...……………...Go to 5
1b. Not an animal…………………………………………………………………….…………………………………………………………………………….…………….Go to 2
2a. Plant that bares fruit or nut………………………………………………….……………………………………………………………………..…………………..Go to 3
2b. Plant that bares only flowers……………………………………………….……………………………………………………………………...……Lonicera iaponica
3a. Plant bares nut………………………………………………………………...……………………………………………………………………..…………….…Quercus abla
3b. Plant bares fruit………………………………………………………………..………………………………………………………………………..…………………..Go to 4
4a. Plant bares yellow fruit when ripe………………………………………….……………………………………………………………….…………Musa acuminate
4b. Plant bares red fruit when ripe……………………………………………..………………….……………………………………...…………Vaccinium Oxycoccus
5a. Vertebrate (backbone)………………………………………………………….……………………………………………..………………………………………….Go to 8
5b. Invertebrate (no backbone)………………………………………………….……………………………………………….………………………………………..Go to 6
6a. Exoskeleton (hard outer covering)……………………………………………………………………………………...…………………………………………..Go to 7
6b. So flesh with shell……………………………………………………………….…………………………………………………...………….………………Helix aspersa
7a. Eight legs……………………………………………………………………………….……………………………………………..………………...….………Euathlus smithi
7b. Six legs……………………………………………………………………………….……………………………………………………....…….……………Oryctes nasicornis
8a. Has fur………………………………………………………………………………..………………………………………………………..………………………Mus musculus
8b. Has feathers……………………………………………………………………….………………………………………..………………………………………………...Go to 9
9a. Long sharp beak…………………………………………………………………….………………………………………….…………………………………………..Go to 10
9b. Short triangular beak…………………………………………………………...…………………………………………….…………………………………………Go to 12
10a. Thick, mul color beak………………………………………………………...…………………………………………………………... ……Ramphastos vitellinus
10b. Thin, needle‐like beak………………………………………………………...………………….…………………………………………………………………..Go to 11
11a. Red colora on on crown……………………………………………..………………………………………………………………………………Dryocopus pileatus 11b. Green colora on on tail………………………………………………….……………………………………………..……………………..Selasphorus platycercus
12a. Gray and orange colora on………………………………………………..……………………………………………..………………………Taeniopygia guƩata
12b. Yellow body colora on……………………………………………………….…………………………………...……………………………..…………Carduelis trisƟs
Figure 3: Dichotomous key.
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Taxonomy
Table 3: Dichotomous Key Results
Organism
i ii iii iv v vi vii viii ix x xi xii xiii
248
Binomial Name
Taxonomy
Post-Lab Questions
1. What do you notice about the options of each step as they go from number one up.
2. How does your answer from question one relate to the Linnaean classification system?
Experiment 2: Classification of Organisms
Materials
Table 2
Figure 4
Procedure
1. Select the first organism from Table 4 (E. coli).
2. Use the “tree” (Figure 4; located at the end of this procedure) start at the base, and answer each question until the organism reaches the end of a “branch”. Write the organisms name in the red box.
3. Repeat this for the remaining organisms.
4. After classification, fill in Table 2 with the correct kingdom for each organism.
Table 4: Key Characteristics of Some Organisms
Organism
Kingdom
Defined Nucleus
E. Coli
Mobile
Cell Wall
Yes
Yes
Yes
No
Yes
Protozoa
Yes
Mushroom
Yes
Sunflower
Yes
No
Bear
Yes
Yes
Photosynthesis
Unicellular
Yes
Yes
Yes
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Taxonomy
Start
Does the organism have a defined nucleus?
Yes
No
Kingdom:
Bacteria
Is the organism mobile?
Yes
Kingdom:
No
No
Fungi
Yes
Kingdom:
Does the organism perform photosynthesis?
Does the organism have specialized cells? Yes
Plant
No
Kingdom:
Kingdom:
Animal
Protist
Figure 4: Experiment 2 - Classification of Organisms Flow Chart
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Taxonomy
Post-Lab Questions
1. Did this series of questions correctly organize each organism? Why or why not?
2. What additional questions would you ask to further categorize the items within the kingdoms (Hint: think about other organisms in the kingdom and what makes them different than the examples used here)?
3. Do you feel that the questions asked were appropriate? What questions would you have asked?
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Lab 17
Bacteria and Archaea
Norris Geyser Basin in Yellowstone Na onal Park
Bacteria and Archaea
Concepts to Explore
•
Identify characteristics of prokaryotes including morphology and energy production
•
Distinguish between prokaryotes that are autotrophic, heterotrophic and chemotrophic
•
Distinguish between prokaryotes based on their morphology including cocci, bacilli, and spirilla
•
Describe the structure of bacterial DNA
Introduction
Bacteria are simple organisms only microns in length, are found everywhere on the planet and are some of the first organisms found in the fossil record. Their varied habitats include soil, acidic hot springs, radioactive waste, seawater, glacial ice and deep in the Earth’s crust.
Bacteria are prokaryotes, cells with no organized nucleus or organelles. Their
DNA is not condensed into chromosomes, but is found as circular loops that float around the cytoplasm. It has been proposed that prokaryotes evolved from endosymbiotic associations after a larger bacterium engulfed a smaller one. This may also explain how mitochondria, chloroplasts, and other organelles developed.
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Did You Know...
Halobacteria is a group of organisms which actually belongs to the archaea domain (not bacteria). Surprisingly, they thrive in concentrated brine nine
Most bacteria are heterotrophic, meaning they cannot manufacture their own times the salinity of sea food. Often they excrete digestive enzymes that decompose the environment water, and remain viable in around them and use the byproducts as energy. However, some are Auto- dry salt crystals for years.
trophic, either obtaining energy from sunlight through photosynthesis, or
Chemoautotrophic, creating energy from inorganic chemicals.
Some prokaryotes have whip-like flagella for locomotion, while others have hair-like pili that allow them to adhere to surfaces. Many bacteria are surrounded by an exterior capsule (coating), which prevents white blood cells (the body’s defense cells) from identifying and destroying them. This coat also makes them stick to target surfaces, and is the reason your teeth feel slimy in the morning!
Bacteria cells exhibit one of several morphologies. The majority are either spherical (Cocci – Figure 1), rod-shaped (Coli – Figure 2), or spiral (Spirillia –
Figure 3).
Many bacteria remain as single cells, while others form colonies, the result of a single bacterium replicating. These colonies assume characteristic shapes
(chains, cluster of “grapes”), which can further classify the bacteria.
Figure 1: Cocci shape.
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Bacteria and Archaea
Bacteria do not undergo mitosis or meiosis like other cells. Each cell replicates its DNA and divides by fission or binary division. This results in identical copies of parent cells (clones). However, genetic variation may be introduced through recombination and mutation.
Small circular segments of DNA, called plasmids, enter the bacteria and integrate into its genetic code. These plasmids often code for genes that make bacteria resistant to antibiotics or viruses. Bacteriophages (often called just phages) are viruses that infect bacteria. Figure 2: Coli shape.
Figure 3: Spirillia shape.
Though we frequently consider bacteria to be harmful, many are not. In a healthy human body, there are 10-times more bacterial cells than human cells. Normally, these microbes are not invasive and do not cause disease (pathogenic). In fact, many benefit to the host by secreting vitamins, digesting food and performing other critical functions. When these bacteria are killed, as is the case with prolonged antibiotic treatment, serious health problems result. The bacteria that live on your skin can also cause disease if they enter the body through a wound or surgical incision (e.g., staph infections that occur after surgery result from staph on your skin getting inside of your body).
Bacteria also play an important role in our environment; binding nitrogen in plants, decomposing and recycling nutrients, synthesizing drugs and making food products (yogurt, sour cream). Bacteria have survived billions of years, in countless environments. Though scientists once thought they would eliminate diseasecausing bacteria, they continue to evolve and develop resistance to even the latest drugs (“anti”-”biotics”).
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Did You Know...
The most prevalent chemical dissolved in the deep ocean waters near hydrothermal vents is hydrogen sulfide. The specialized bacteria that live near these vents use hydrogen sulfide as their energy source, instead of sunlight (chemoautotrophs).
The bacteria, in turn, sustain larger organisms in the surrounding area. The clams, mussels, tube worms, and other creatures at the vent have a symbiotic relationship with bacteria. They could not live without each other! The giant tube worms have no mouth or gut to digest food. Instead, the worm depends on the bacteria that have invaded its body to survive. There are around 285 billion bacteria per ounce of tissue within this tube worm. The plumes at the top of the worm's body are filled with blood, which contains hemoglobin that binds hydrogen sulfide and transports it to the bacteria that lives within the tissue. In return, the bacteria oxidize the hydrogen sulfide and convert carbon dioxide into carbon compounds that nourish the worm.
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Bacteria and Archaea
Pre-Lab Questions
1. Based on the scientific name Streptococcus agalactiae, what morphology would you expect these cells to have? 2. Name at least three animal structures that would be analogous to bacterial flagella.
3. Hypothesize how over-washing of hands can affect the population of “good” bacteria that resides on the human skin.
Experiment 1: Testing the Environment
Bacteria is found in almost all environments. Most are harmless, but there are those that cause disease. In this experiment, you will be test different environments for the presence of bacteria. You may be amazed to see how many different species are present on surfaces you come in contact with every day!
Materials
30 mL 8.25% Bleach Solution
*Hot Pad
4 Cotton Swabs (Sterile)
*Microwave or Boiling Water Bath
125 mL Nutrient Agar
*Refrigerator
60 cm Parafilm®
*Scissors (to cut the Parafilm®)
Permanent Marker
(4) 9 cm Petri Dishes
*You Must Provide
Notes About Working With Agar Plates…
•
Prepared agar dishes should be stored upside-down in the refrigerator until used. This will prevent condensation from disrupting the growing surface.
•
After inoculating, replace the cover on the dish, seal with Parafilm®, and store upside-down in a warm location (not to exceed 37.7 °C or 100 °F).
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Bacteria and Archaea
•
You should see growth within a few days. The plate will start to smell once microorganisms are growing. •
Before disposing plates, kill the microorganisms by pouring bleach solution onto the agar surfaces and let sit for 20 minutes.
Procedure
1. Loosen or remove the cap on the Nutrient agar bottle. Place in the microwave (if you do not have a microwave, place the bottle in a heat-safe bowl and pour boiling water around the bottle) and heat until the entire bottle of agar is liquefied. You will need to remove the bottle and swirl every 10 seconds to distribute the heat.
2. If you notice the liquid boiling over, STOP the microwave and let the bottle cool down before handling.
With a hot pad protecting your hands, remove the bottle from the microwave. Use caution when removing the bottle from the microwave as it will be HOT!
3. Gently swirl the bottle to mix the solution. Pour enough of the liquefied agar solution into the bottom half of 4 petri dishes so that it covers the entire bottom of the dish. Place the lids onto the dishes and CAREFULLY transport these plates into a refrigerator. Allow the plates to sit for 24 hours.
4. After 24 hours, remove the four agar plates from the refrigerator and allow them to sit at room temperature for at least one hour.
5. After the plates have warmed to room temperature, locate a surface to swab for bacteria. Things such as shoes, a table, teeth, bathroom doors, and shopping carts can be great sources. Try to avoid surfaces that are frequently cleaned with antibacterial detergents and soaps. On the bottom of the plate, write the name of the surface to be swabbed with your permanent marker.
6. Remove the lid from the agar plate. Unwrap one sterile cotton swab and smear the surface, being sure to roll the cotton swab to cover all sides.
7. Carefully streak the cotton swab onto the surface of the gelatinous medium (agar), being sure to start at the top and work down in a zigzag motion.
8. Place the lid onto the agar plate and seal it with a strip of Parafilm (hold one end of the Parafilm firmly against the side of the petri dish and stretch the other side to cover the entire perimeter). Place the plate upside down in a warm area to incubate.
9. Repeat this for two additional surface areas and agar plates.
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Bacteria and Archaea
10. For the last plate, label it “control.” Do not rub your sterile cotton swab on any surface, but rather take it straight from the package and streak it onto the plate. This will test for your accuracy in keeping the plates and swabs sterile while performing the experiment.
11. Let the plates incubate in a warm area for three days and then observe the growth. Answer the questions below.
12. After the experiment and after answering questions 1 - 3, set aside the plate with the most bacterial growth for the next experiment. Pour the bleach solution onto the surface of the remaining agar plates, allowing it to cover the entire surface. Let the plates sit untouched for 10 - 20 minutes. Then, seal plates with Parafilm® and dispose appropriately.
Post-Lab Questions
1. In the space provided below, draw your plates and indicate the number of different colonies and identify the colony shape.
2. Which plate grew the most bacterial species? Was this a surprise? Why or why not?
3. Was your control plate free of bacterial colonies? If not, how do you think the swabs were contaminated?
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Bacteria and Archaea
Experiment 2: Measuring Antibiotic Resistance
In this experiment you will look at the ability of common antibiotics and other anti-bacterial agents to kill bacteria commonly found in your environment. You will use the bacterial lawn from the plate that produced the largest quantity of bacteria in the first lab.
Materials
1 Ampicillin Disc (marked)
1 Penicillin disc (marked)
10 mL 8.25% Bleach Solution
Permanent Marker
Forceps
Ruler
1 Kanamycin Disc (marked)
Parafilm®
1 Nutrient agar plate from the previous lab
Note: Be sure to wear gloves and use the forceps when handling the discs. Always pick them up if spilled.
Procedure
1. Using one of the plates covered with bacteria from the first lab, draw four even quadrants onto the bottom of the plate. Label them penicillin, ampicillin, kanamycin and control.
2. Wearing your gloves, goggles, and apron, open the plate and place the appropriate disks into the appropriate quadrants using forceps. Do not place anything in the control quadrant. DO NOT TOUCH THE
PLATE WITHOUT GLOVES.
3. Place the lid back onto the plate and cover it with Parafilm®. Allow it to incubate in a warm area for 3 days. 4. After 3 days, observe the zone of resistance from each disc (the region surrounding the disks where no bacteria grew). Using a ruler, measure these zones from the bottom of the agar plate.
5. After you are done with the experiment, pour enough bleach solution to cover the surface of the agar. Allow the plate to rest for 10 - 20 minutes. Seal the plate with Parafilm® and dispose appropriately.
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Bacteria and Archaea
Post-Lab Questions
1. Did your bacterial lawn contain only one species of bacteria? If not, why do you think that is? Can you tell?
2. Which antibiotic was most effective in killing the bacterial lawn? Which was the least effective?
3. Each bacterial species shows different antibiotic susceptibility. What can you say about the bacteria that you grew?
261
Lab 18
Protista
Pictured Above: Amoeba
Protista
Learning Objectives
•
Describe the diversity and complexity of organisms classified under Protista
•
Compare and contrast protist motility including pseudopodia, cilia, and flagella
•
Distinguish between protists that are autotrophic, heterotrophic, and mixotrophic
•
Distinguish between filamentous forming and colony associating protists
Introduction
Protists are some of the oldest and most diverse organisms on the planet. Most species which are grouped within the kingdom Protista share very few characteristics except for being eukaryotic. Some researchers refer to the kingdom Protista as a “catch-all” for organisms that do not fit into any other kingdom. Protists may be unicellular, multicellular or colonial; mobile or non-mobile; and possess mitochondria or chloroplasts.
Some protists are ever visible with the naked eye, although most require magnification.
Most protists reside in some type of water, whether it be the ocean, a pond, or within the human body. Their energy production can be either autotrophic or heterotrophic, with some switching between the two
(mixotrophic). All protists can reproduce asexually and some have the ability to reproduce sexually.
Protists can be defined as being unicellular, multicellular filamentous forming, or multicellular colony forming.
Chlamydomonas is a unicellular protist, more commonly known as green algae. There are over 600 species of this protist, but all contain chloroplasts which allow them to convert solar energy to chemical energy and provides them with their characteristic green color (Figure 1).
Figure 1: Chlamydomonas (green algae) are often found in stagnant water or damp soil.
265
Protista
Protist Locomotion
Protists vary significantly in their motility. Some are entirely stationary, while others employ either flagella, cilia or pseudopodial movement. Flagella are common throughout the kingdom Protista, although they are also used by some prokaryotes. They are tail-like extensions that rotate or whip back and forth to produce locomotion. Flagellar movement may be due to a stimulus, such as a chemical gradient or food source. Depending on their location, flagella can either push or pull the protists through liquid medium. Some protists may have a single flagellum, while others have Figure 2: Giardia have two nuclei and four flagella. multiple flagella, like Giardia.
Ciliated protists use cilia, short, hair-like protrusions that enable the organism to move. The uniform motion of the cilia creates a “wave” that propels the protist. This wave also serves as a means for feeding by sweeping the food to them. One example of a ciliated protist is a paramecium. Other protists, such as amoebas use psuedopodial movement by extending a temporary protrusion called a pseudopod (Greek for “false foot”) to slowly crawl forward. Psuedopods are also used in feeding.
Slime Molds, Algae, and Protozoans Figure 3: Paramecium move using a coordi- The diversity of protists is vast, but this lab focuses on three prominent protists: slime molds, algae and protozoans. nated waving motion of their cilia.
Slime-Molds
Slime molds, often considered the link to the kingdom Fungi, share similarities with the kingdom Fungi such as appearance, nutrient acquisition, and the ability to form spores. In fact, some researchers refer to slime molds as “Fungus-Like Protists”. However, they differ in many key characteristics such as reproductive technique, life cycle and cellular make-up. Slime molds are phagocytic heterotrophs, meaning they surround and engulf particles with cytoplasm protrusions. They possess a complex reproductive cycle which includes mobile component (similar to that of animals), and an immobile component (similar to that of plants) when spore formation occurs.
Slime molds are either plasmodial or cellular. Plasmodial slime molds (Figure 4) are large, single-celled organisms with numerous nuclei. These nuclei are formed when many cells come together and fuse. Slime molds can sometimes be described as brightly colored cytoplasmic “blobs” which feed off of other microorganisms, such as bacteria. They can move via amoeboid movement, employing a method which mimics the pseudopodial movement of amoebas on a larger scale. They are quite useful in scientific studies because of their size.
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Protista
Cellular slime molds spend most of their life as individual single cells, but can come together and form a large multicellular organism. As the cells join, a slime covers the organism, allowing for uniform movement. Though the cells are attached, they do not fuse and remain distinct. They provide a useful tool to examine the formation of multicellular organisms and the ability of cells to recognize one another.
Algal Protists
With a multitude of colors, algae are often the most recognizable member of the kingdom Protista. Algae are photosynthetic autotrophs, meaning they possess chlorophyll and often other photosynthetic pigments. A select few are also heterotrophs, making them a mixotroph (an organism that has the ability to be photosynthetic as well as heterotrophic). There are three main types of algae: green algae, red algae and brown algae, each of which is distinct.
Figure 4: Plasmoidal slime molds typically span approximately three centimeters, but some have been documented to measure several meters in diameter!
Algae serve a number of important commercial functions. They produce over half of the oxygen in the
Earth’s atmosphere and are used for a number of commercial applications. For example, algae can be used to create ice cream, shaving cream, air fresheners, laxatives, dyes, pastry fillings, syrups and more).
Protozoans
Protozoans are typically unicellular organisms which require magnification to be seen. Like slime molds, they are phagocytic heterotrophs that surround and engulf small organisms. Since protozoans are motile, using psuedopodial movement, flagella or cilia, they are found in multiple environments. Though not all protozoans are harmful, as a group they are responsible for more disease and sickness throughout the world than any other group of organisms. Some of these diseases include Malaria (caused by the plasmodium protozoan), Giardia (caused by the giardia lamblia protozoan), African Sleeping Sickness (caused by the trypanosome brucei gambiense protozoan) and amoebic dysentery, also known as “Montezuma's Revenge” (caused by the entamoeba histolytica protozoan).
267
Protista
Pre-Lab Questions
1. Hypothesize what type of environments make being autotrophic, heterotrophic or mixotrophic an advantage to a protist.
2. Many algae are photosynthetic like plants. Why then are they not classified as such?
3.Some protists are colony forming. Hypothesize why this may be an advantage or disadvantage.
Experiment 1: Viewing Preserved Species of Protists
In this lab you will look at preserved specimen slides of the Protist kingdom. While examining the slides, pay close attention to the individual characteristics of each organism (i.e. flagella, color, etc.).
Materials
Euglena Digital Slide Images
Paramecium Digital Slide Images
Procedure
1. Observe the detail of each organism (does it have flagella, what size is it, etc.). Look at Question 1. Draw each organism and label the major structures.
Post-Lab Questions
1. For each digital image (Figure 5 and 6), label the major structures you can see. What type of locomotion do you think this protist employs?
2. What advantages do you think a protist would gain by having a unicellular, filamentous, or colonial form?
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Protista
Figure 5: A digital slide picture of Euglena 1000X
Figure 6: A digital slide picture of Paramecium 1000x
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Protista
Experiment 2: Viewing Live Specimens
While observing the protists, you will notice that each member will exhibit different types of locomotion, speed, etc. You will observe a slowing reagent in part of the lab to get a better look at the fast moving protists.
Materials
Live protist specimen videos on the eScience Labs’ Student Portal
Procedure
1. Observe the movement of the protists with and without slowing agent. Notice the different types of locomotion employed by different types of protists.
Post-Lab Questions
1. In the space provided below, draw the various members of the protist kingdom that you were able to observe. Next to each drawing, describe the speed and type of locomotion you observed.
No slowing reagent
With slowing reagent
2. Did you notice a difference in the protists when the slowing agent was used? Which members were you able to see more clearly? Were there species that were still moving too fast to see clearly? Could you identify any species specifically? Do any of them look similar to specimens you observed in the first lab?
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Lab 19
Fungi
Fungi
Learning Objectives
•
Distinguish characteristics between the four major phyla of the fungi kingdom
•
Connect decomposers and saprophytes including hyphae and mycelium
•
Hypothesize why fungi have different and complex reproduction techniques including spores and dikaryon •
Describe the commercial uses of fungi
Introduction
Fungi are immobile, multicellular organisms (yeast is unicellular) that have some unique characteristics and play a profound role in ecology and human health. Fungi fall into four phylums:
•
Zygomycota (bread molds)
•
Ascomycota (sac- yeast and molds)
•
Basidiomycota (club mushrooms, rusts)
•
Deuteromycota (no identifiable sexual stage)
Fungi are decomposers that secrete enzymes to break down material into soluble organic compounds, which they can then absorb. They are saprophytes that digest dead and decaying material (e.g. ground, rotting wood, dung) and are critical in decomposing and recycling nutrients for the environment.
Multicellular fungi are primarily filamentous, with long, thin, somatic cells called hyphae. Unlike most plant and animal cells, hyphae can have more than one nucleus. Mycelium, a network of hyphae that lack distinct cells, are the structures that invade and digest material in soil, wood and other matter. Chitin is a light but strong substance found in the cell wall (in hard shells of insects and crustaceans) that adds rigidity and strength. Since the cytoplasms of hyphae are linked within the mycelium, a unique form of mitosis occurs in fungi. Instead of dividing nuclear material and the cytoplasm, mitosis occurs in the nucleus, which then divides (the nuclear envelope does not dissolve).
Reproduction
Spores are the basic reproductive structure of fungi. Fungi reproduce either asexually or sexually, depending on their environment. During the asexual phase, microscopic haploid spores are created by mitosis (the spores are identical). When they land in the right conditions, they germinate and grow new hyphae. There are five types of asexually produced spores:
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Fungi
1. Arthrospores, produced in fungi that have divided hyphae, are individual segment of the hyphae that break off into spores.
Figure 1
2. Chlamydospores, a segment of the mycelium, become thick-walled and breaks away as the spore. Figure 2
3. Sporangiospores are produced in a sac-like structure called the sporangium.
274
Figure 3
Fungi
4. Conidiospores are not enclosed, but are produced in the conidiophores.
Figure 4
5. Blastospores are produced by a budding process (mitosis with unequal division of the cytoplasm).
Figure 5
Fungi form gametes through mitosis (unlike other multicelled organisms) and spend the majority of their life cycle in a haploid state (1n).
During sexual reproduction, hyphae of two different specimens, from the same species, meet and fuse together to form a dikaryotic cell (1n+1n). Once the nuclei have fused into a zygote, it goes through meiosis producing that mature into a new haploid organism.
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Fungi
Fungi produce three types of spores:
1. The fusion of two nuclei produces a thick-walled zygosporangium which manufactures “+” and “–“ haploid zygospores
Figure 6
2. Two fungi touch exchange nuclei that fuse together in a sac-like structure called the ascus. Meiosis then produces four nuclei which subsequently undergo mitosis, producing eight ascospores.
Figure 7
276
Fungi
3. Mycelia fuse together to form a secondary structure with two nuclei. They maintain this dikaryotic state until fruiting occurs and the “gills” of the mushroom develop. Meiosis then produces four nuclei which migrate into the gills forming basidiospores.
Figure 8
Many fungi are harvested commercially for food, bread leavening, alcohol production, antibiotic production and other uses. Many symbiotic relationships have evolved between fungi and other plants and animals.
Fungi can produce chemicals that make food unpleasant, carcinogenic, or poisonous. They can cause athlete’s foot, ringworm and can kill plants (e.g. potatoes, elms and oak trees). Some are parasitic, feeding off living cells, absorbing their nutrients and killing their host.
Figure 9: Mushroom life cycle
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Fungi
Definitions:
1. Chitin: A resilient and tough polysaccharide found in the cell wall of fungi, as well as the exoskeletons of insects and crustaceans
2. Dikaryon: Two cells that merge into one without fusing nuclei; a cell with two haploid nuclei
3. Hyphae: The branching, filamentous cells of fungi
4. Saphrophyte: An organism that feed on non-living organic matter
Pre-Lab Questions
1. Hypothesize why a fungus would use spores as a mechanism for reproduction.
2. How might the environment affect a fungus reproducing sexually or asexually?
3. What characteristics are more plant-like? Animal-like?
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Fungi
Experiment 1: Growing and Observing Zygomycota (Bread Mold)
Bread mold will generally ruin a day when you are looking forward to a sandwich, but will make it much easier to observe a live fungus. In this lab you will be cultivating the common bread mold. Any type of bread can be used, but fresh bakery bread will likely grow more colorful species since it does not contain the preservatives that many processed varieties do.
Materials
Permanent Marker
*2 Pieces of White Bread
Spray Bottle Lid and Empty Bottle
*Water
2 Resealable Bags
Rhizopus Digital Slide Images
* You Must Provide
Procedure
1. Use the permanent marker to label each of the two resealable bags as “Wet” or “Dry”.
2. Take one slice of bread and, spray it with water until moist. Avoid soaking the bread.
3. Place the moistened bread in the resealable bag labeled “Wet” and seal.
4. Then, place the second piece of bread in the resealable bag labeled “Dry” and seal.
5. Create a hypothesis regarding how the water will affect the bread over time. Be sure to address which piece of bread you think will host more growth, and why. Include your hypothesis in Post-Lab Question 1.
6. Incubate the bags for 3 - 7 days (depending on how fresh the bread is) in a dark, warm spot.
7. Examine the bread once mold starts growing.
8. Examine the digital slide pictures of Rhizopus (Figure 10) and identify the structures.
Figure 10: Rhizopus digital slide pictures; 100X (above left), 400X (above right).
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Figure 10 (continued): 1000X
Post-Lab Questions
1. Include your hypothesis from Step 5 here.
2. What structures did you see in the bread mold?
3. Why was it important to moisten the bread before sealing it in the resealable bag?
4. What type of control (positive or negative) did the dry bread provide? Explain why.
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5. Is the bread mold reproducing sexually or asexually? How could you determine this?
6. What would you expect to find if you left the bread for six months?
7. Were multi-nucleated hyphae prevalent when looking at the Rhizopus slide? How do you know?
Experiment 2: Observing Ascomycota (Yeast)
Yeast is a commercially important member of the Fungi Kingdom. It leavens bread and produces beer and wine. Yeast is unique because it is unicellular and reproduces by mitosis and budding.
Materials
Yeast Digital Slide Images
Peziza Digital Slide Images
Procedure
1. View the digital slide image of yeast (Figure 11). Locate the budding cells and label what you see in the space below. Using the slide as a reference, draw a budding yeast cell and include important structures
(ascus, ascospore) on your picture.
2. View the digital slide image of Peziza (Figure 12). Identify the asocarp, asci, ascospores, and sterile hyphae.
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Figure 11: Yeast 1000X; field 1 (left) and field 2 (right)
Figure 12: Digital slide images of Peziza; 100X (above left), 400X (above right); 1000X (below)
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Post-Lab Questions
1. Compare yeast to the Peziza slide. Identify at least one difference, and one similarity.
2. How many ascospores are in each ascus of the yeast?
3. Are the ascospores of the Peziza inside or outside of the asci?
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Lab 20
Energy and Photosynthesis
Energy and Photosynthesis
Learning Objectives
•
Identify the four principle structures of a chloroplast: thylakoids, grana, lamella and stroma
•
Use photosynthesis to explain how the sun affects all organisms
•
Compare and contrast the light-dependent and light-independent reactions that occur during photosynthesis
Introduction
Photosynthesis is the process by which most plants, certain bacteria, and some algae harness light energy and manufacture food. In other words, it uses solar energy (light) to produce chemical energy
(carbohydrates). Photosynthesis consumes carbon dioxide (CO2, a greenhouse gas) and releases oxygen
(O2) (Figure 1). Nearly all life on Earth relies on this reaction in some format. Although there are many variants of photosynthesis found throughout nature, one of the most common products formed through photosynthesis is a sugar molecule called glucose (C6H12O6).
Figure 1: C6 Photosynthesis.
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All photosynthetic reactions are either light-dependent or light-independent. Both types occur in specialized organelles called chloroplasts in eukaryotes (plants and some algae species). The proteins involved in photosynthesis in photosynthetic bacteria are located in the cell membrane of these prokaryotes.
Chloroplasts, similar to that of mitochondria, are thought to have originated through an endosymbiotic event. That is, at some point in eukaryotic evolution, photosynthesizing bacteria (specifically cyanobacteria) were engulfed and became permanently associated with the eukaryote. In fact, chloroplasts maintain their own, unique circular double stranded DNA.
A chloroplast has four principal structures that play a role in photosynthesis (Figure 2):
Figure 2: Chlorophyll gives plants their green color. Wheat grass juice, common in health food stores, has a pronounced dark green color due to its high chlorophyll content.
1. Thylakoids: Small disk-shaped structures. These contain a pigment called chlorophyll which captures light energy.
2. Grana: Stacks of connected thylakoids.
3. Lamellas: Structures that link grana together.
4. Stroma: The fluid within the chloroplast, similar to cytoplasm. This fluid surrounds the grana and contains the liquids and molecules required for the Calvin Cycle. Chloroplast DNA and ribosomes are also located within the stroma.
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Plant Pigments
Plants have a number of different pigments which give plants their color and absorb light. Chlorophylls and carotenoids are two important classes of pigments. Chlorophylls are the greenish pigments and include several kinds.
Chlorophyll A is found in all photosynthesizing plants, algae and cyanobacteria. Chlorophyll B and chlorophyll C are less widespread and are only found in certain groups of organisms.
Carotenoids include the red, orange and yellow pigments. Carotenoids are sometimes referred to as accessory pigments because they pass the absorbed energy to chlorophyll when they absorb sunlight. Hundreds of known carotenoids are categorized into two major divisions: carotenes and xanthophylls. Beta carotene is the most abundant carotene in plants. This orange pigment gives carrots their color. Xanthophyll are yellow pigments and, similar to carotenes, are found in the leaves of most green plants.
Oxidation-Reduction Reactions (Review)
?
Did You Know...
Most of us have heard of the circadian cycles that humans engage in during sleep. But most people don’t know that circadian cycles also affect plant species to help them sense ambient light levels.
This cycle is regulated by oscillating biochemical processes which promote or inhibit the production of proteins involved in photosynthesis.
This regulation is critical to a plant’s ability to maintain its base anabolic actions which rely on solar energy.
Oxidation-reduction reactions, commonly referred to as redox reactions, are involved in photosynthesis. Redox reactions always include two separate reactions: an oxidation reaction and a reduction reaction. These reactions are so intricately connected that each oxidation and reduction reaction is often considered a halfreaction. Electrons are lost in oxidation reactions, and the same electrons are gained in the associated reduction reaction as the molecule transitions from the reactant to the product. When a molecule or atom undergoes oxidation, it is said to have been oxidized. Oxidizing agents are responsible for oxidation. Once the electrons are lost, the molecule is considered a cation. Cations are molecular ions which have a positive charge. A reduction reaction is one in which a molecule gains electrons as it transitions from the reactant to the product. Once the electrons are gained, the molecule is considered an anion. When a molecule or atom undergoes reduction, it is said to have been reduced. Reducing agents are responsible for reduction. Anions are molecular ions which have a negative charge.
Light-Dependent Reactions
Light-dependent reactions take place in chlorophyll A, chlorophyll B, and carotenoid pigments. Electrons act as the energy carrier throughout these reactions. Operating in tandem with other pigments, chlorophyll breaks the chemical bond of water and releases hydrogen and oxygen. These products, along with carbon dioxide undergo additional modifications in the stroma during the light-independent reaction stage (also known as the
Calvin-Benson Cycle). This ultimately results in the production of glucose, which can then be used for energy.
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Wavelength vs. Percent Absorbed of Different Plant Pigments
Figure 3: Wavelength vs. Percent Absorbed line graph. Violet: 400 nm, Blue: 475 nm, Green: 515 nm,
Yellow: 570 nm, Orange: 620 nm, Red: 675 nm.
Photosynthesis: White Light and Green Light
White light is a form of solar energy that contains all the wavelengths in the spectrum (i.e. “colors”: blue, green, yellow, etc.). Each pigment in a plant absorbs specific wavelengths of light, with most plants being unable to absorb green wavelengths. These are reflected back, which is why plants typically appear green. All of the other wavelengths (colors) are absorbed by the plant and initiate the formation of chemical energy. Figure 3 illustrates how each pigment absorbs specific wavelengths. Remember, color corresponds with the wavelength. Light-Independent Reactions (The Calvin-Benson Cycle):
After the light-dependent reaction, the light-independent reaction takes resultant products and converts them to carbohydrates and oxygen in a process known as the Calvin-Benson Cycle (Figure 4). The CalvinBenson Cycle is composed of multiple reactions that link carbon atoms to form simple sugar molecules such as glucose. This cycle begins when the enzyme RuBisCO (ribulose 1,5-bisphosphate carboxylase/ oxygenase) acquires carbon from the atmosphere in the form of carbon dioxide. Carbon fixation, reduction, and regeneration of ribulose then proceed to complete the Calvin-Benson Cycle.
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Figure 4: The Calvin-Benson Cycle.
Chromatography
Chromatography is a set of techniques that can physically separate components of a mixture (analyte) allowing for purification and analysis of these components. The physical and chemical properties of each analyte affect the rate of migration. For example, analytes with low solubility will migrate a shorter distance than analytes with higher solubility. These different rates, consequently, separate the analytes. In paper chromatography, the analytes are separated along a piece of chromatography paper.
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Paper chromatography is used to separate analytes, such as plant pigments, along a piece of chromatography paper. When the end of a piece of paper is dipped into water, the water molecules find new molecules to bond. The water molecules climb up the paper being replaced by new water molecules below. Solutes, which might be dissolved in the water, will also be carried along up the paper. The different solutes are carried along at different rates because they are not equally soluble. They are also differentially attracted to the fibers of the paper through the formation of intermolecular bonds, such as hydrogen bonds. Plant pigments can be separated by paper chromatography. After running the chromatogram, a retention factor (Rf factor) can be calculated for each analyte. The Rf is a ratio of the distance the analytes travel relative to the distance the eluting solvent travels.
The different pigments found in plants can be separated using paper chromatography. Beta carotene is able to travel relatively far up the paper because the pigment does not form any hydrogen bonds with the paper.
Xanthophyll, on the other hand, contains an oxygen group which forms hydrogen bonds with the paper.
Therefore, this pigment will be seen lower than beta carotene. Chlorophyll contains both oxygen and nitrogen, which bind even stronger to the paper and will travel even less than xanthophyll when separated with paper chromatography. You will be using pH indicators to detect carbon fixation, or the depletion of carbon dioxide, during the proceeding lab experiments. The concentration of carbon dioxide molecules decreases as carbon dioxide is reduced and oxygen is released. This causes the environmental pH to rise and the solution to become more basic. Pre-Lab Questions
1. Describe how the functional units for beta carotene, xanthophyll, chlorophyll A, and chlorophyll B are different. Be sure to identify the subunits that adhere to paper during chromatography.
2. Describe a technique for measuring photosynthetic rate.
3. Many deciduous trees have leaves which turn yellow in the fall. What do you suppose is happening in the leaves at the cellular and molecular level?
4. Chloroplasts and mitochondria are both are unusual in that they have double membranes and contain their own set of DNA. Can you think of any explanations for this observation?
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Experiment 1: Paper Chromatography
In this experiment, you will separate plant pigments using chromatography. You will also measure the rate of photosynthesis in isolated chloroplasts using the reduction of the dye 2,6-Dichloroindophenol (DPIP) as the measurement tool. The transfer of electrons during photosynthesis reduces DPIP, changing it from blue to clear. Materials
10 mL 4.5% Acetic Acid (Vinegar), C2H4O2
100 mL 0.5 M Sucrose Solution (cold!), C12H22O11
10 mL Acetone (Nail Polish Remover), C3H6O
3 Test Tubes (Glass)
30 cm Aluminum Foil
Test Tube Rack
(4) 100 mL Beakers
*Cutting Board
(1) 250 mL Beaker
*Kitchen Knife
20 cm Cheesecloth
*Light Source
3 mL 1% 2,6-Dichloroindophenol, DPIP
*Pencil
(1) 12 x 12 cm Chromatography Paper Piece
*Quarter
(1) 100 mL Graduated Cylinder
*Scissors
10 mL Mineral Oil
**Spinach Leaves (Fresh)
3 mL 0.1 M Phosphate Buffer, P
*Tape (masking or Scotch®)
8 Pipettes
*Water
(1) Resealable Plastic Bag
Rubber Band (Large; contain latex, handle with
*You Must Provide
gloves on if allergic)
*Keep these leaves if performing Experiment 2
Ruler
within a few days of Experiment 1.
Wooden Stir Stick
Procedure
Part 1: Paper Chromatography
1. Place the bottle containing 100 mL of sucrose solution in the refrigerator. Allow the solution to rest here for the remainder of Part 1; it will be required in Part 2.
2. Use the permanent market to label each 100 mL beaker as 1, 2, 3, or and 4.
3. Add 10 mL water to Beaker 1, 10 mL acetone to Beaker 2, 10 mL mineral oil to Beaker 3, and 10 mL acetic acid to Beaker 4. Cover the beakers with aluminum foil, and set them aside.
4. Use a pencil to draw a line approximately one cm from the bottom (across the bottom edge) of each piece
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Energy and Photosynthesis of chromatography paper. Then, carefully cut each piece in half so that you have four, pieces of identical size (each piece should have the pencil line going across the bottom edge of the paper).
5. Use a quarter to extract pigment from a fresh spinach leaf by placing the leaf in between the filter paper and the coin. Firmly rub the edge of the quarter back and forth over the pencil line drawn in Step 2. Use a fresh section of the leaf for each rubbing (Figure 5).
Figure 5
6. Place one piece of filter paper in Beaker 1, with the pigment-side towards the bottom of the beaker. Tape the filter paper to the beaker so that the bottom of the filter paper is submerged in the solvent, but the pigment line is not. DO NOT ALLOW THE PIGMENT TO TOUCH THE SOLVENT!
7. Monitor the set-up as the solvent travels up the paper. Remove the paper when the solvent is about 1 cm from the top of the paper. Immediately mark the solvent front and the location of each analyte band.
8. Measure the distance from the original pencil line to the solvent front, and the original pencil line to each of the pigments. Record the data in Table 1.
9. Calculate the Rf using the following equation:
10. Repeat Steps 6 – 9 for Beakers 2, 3, and 4. Be sure to use a new piece of filter paper for each beaker.
Rf = distance pigment migrated (mm) distance solvent migrated (mm)
Part 2: Chloroplast Isolation
11. Obtain a knife and cutting board. Carefully cut across a few spinach leaves to isolate chloroplasts from the leaves. 12. Place the leaves in a resealable bag. Measure and pour 100 mL of the cold sucrose solution (prepared in
Part 1: Step 1) into the resealable bag with the spinach leaves.
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13. Remove any air from the plastic bag and seal completely. Mash the solution for 2 minutes.
14. Lay a piece of cheesecloth over a 250 mL beaker, and push the cheesecloth into the beaker approximately four to five cm with your finger. Secure with a rubber band.
15. Slowly pour the spinach/sucrose solution into the cheesecloth. Allow the cheesecloth to filter the solution for several minutes; or, until all of the liquid has passed through the cheesecloth.
16. Once the liquid has drained, discard the collected solids. You may wish to squeeze the cheesecloth to extract any remaining liquid before throwing it away if liquid appears to be caught.
Part 3: Photosynthesis
17. Use the permanent marker to label three test tubes as 1, 2, and 3. Place the tubes in the test tube rack.
18. Add 3 mL water, 1 mL phosphate buffer, and 1 mL DPIP to each of the three test tubes.
19. Use the wooden stir stick to swirl the chloroplast solution
(prepared in Part 2). Then, add two drops of the solution to Tubes 2 and 3.
20. Immediately cover Tube 2 with aluminum foil.
21. Place Tube 1 and Tube 3 in a sunny location or under a strong light. If you place the tubes in front of a light, fill a beaker or clear glass with water and position in between the light source and the test tubes as shown in Figure 6.
Figure 6: A heat-sink for test tubes placed in front of a light source.
22. Monitor the test tubes for several hours, and record how long it takes each tube to turn from blue to clear. After the tubes exposed to light turn clear, remove the aluminum foil from Tube 2 and immediately note the color. Record your data in Table 2.
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Table 1: Part 1: Chromatography Data
Solvent
Distance from Original Line to Solvent Front
Number of Bands
Rf Factor
Acetic Acid
Acetone
Mineral Oil
Water
Table 2: Part 3 - Photosynthesis Data
Test Tube
Time Required to Change Color
1
2
3
Post-Lab Questions
1. What did the different colored bands signify in each solvent for Part 1? What pigments can you associate them with?
2. What is the osmolarity fluid used in Part 2? Why is this important? Why is it essential to keep it cool?
3. How could you modify this experiment to show the effects of different wavelengths of light on the photosynthetic rate?
4. Some plants (grasses) tend to contain a greater concentration of chlorophyll than others (pines). Can you develop a hypothesis to explain this? Would it be testable?
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Lab 21
Plant Circulation
Plant Circulation
Learning Objectives
•
Compare and contrast the structure and function of xylem and phloem in plant circulation
•
Hypothesize what factors affect transpiration rate
•
Explain the role water plays in transpiration
•
Compare and contrast transpiration with non-vascular water consumption
Introduction
Plants are multi-cellular, eukaryotes. They require energy and nutrients to engage in vital cell processes.
However, unlike animals which have a heart and blood to circulate critical elements, vascular plants must rely on their roots to uptake water and minerals from soil, and a specialized circulatory system to transport the nutrients to all other parts of the plant. This system is composed of xylem and phloem (Figure 1).
Xylem and Phloem
Xylem are hollow vessels produced from dead cells. They have additional support in their cell walls, which allows them to withstand the heavy weight imposed by the overall plant structure. Water and nutrients are transported from roots to leaves through the xylem conduits. Phloem are composed of living cells which form long, sieve-like tubes to transport the carbohydrate products of photosynthesis from the leaves to the rest of the plant. This process is called translocation. The vascular system allows plants to grow in diverse climates and to an extreme size. For example, Sequoia trees can live up to 2,200 years and grow to a height of 379 feet! That’s longer than a football field!
Figure 1: Root cross-section.
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Figure 2: Transpiration.
Transpiration
Water moves up the xylem, against the force of gravity. Water which reaches the surface of the leaves through the xylem may then evaporate and diffuse out of the stomata into the atmosphere in a process called transpiration. Stomata are small pore like objects which regulate the flow of water by opening to release water into the atmosphere through evaporation. There are millions of stomata in a single plant leaf.
When water evaporates, additional water and nutrients must be withdrawn from the ground to replace the water which has been lost. This process begins when water is absorbed by root cells and flows up the stem to the leaves and flowers, where it is allowed to escape (Figure 2). In this way, a continuous process of
“drinking” water from the soil and evaporation takes place.
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Figure 3: Evaporation process flow.
Transpiration Rate
A few key factors contribute to transpiration rate. These factors include humidity, light, temperature, and wind.
Humidity is inversely related to transpiration rate; in other words, increased humidity decreases the rate of transpiration. Light, temperature, and wind are all directly related to transpiration rate. For example, bright, windy, and high temperature conditions may all increase the transpiration rate.
Chemical and Physical Properties of Water
The chemical formula of water is recognized as H2O. Looking at the chemistry of the molecule, you can see that two hydrogen atoms are bound to one oxygen atom (Figure 4). However, the hydrogen atoms do not bind equidistant from each other.
Instead, two hydrogen atoms bond to the same side of the oxygen atom, creating molecular polarity. In other words, there is a positive charge on one side of the molecule, and a negative charge on the other. Since opposite charges attract, water molecules are attracted to other water molecules. In this form, or as a solid (ice), or gas (water vapor), water serves a great Figure 4: A molecule of water (red = oxygen atom; blue = hydrogen atoms). many purposes in the Earth’s atmosphere.
Due to the cohesive nature of water (polarity), as a water molecule leaves the plant, another takes its place.
The surface tension among the water molecules in the xylem, all the way down to the roots, ensures that as one molecule is lost to evaporation another replaces it in the roots. Think of sucking on a straw, as you suck liquid at the top, it is replaced by liquid at the bottom.
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Non-vascular plants (bryophytes) do not have a specialized circulatory system and are growth-restricted based on the availability of water. These low-growing plants, such as moss and liverwort species, usually thrive in dark, damp habitats where they can absorb water from their surface. Although they lack vascular tissues, true roots, leaves, and stems, they can transport water internally. Like sponges, they must imbibe the water needed for survival from their surrounding atmosphere through processes such as diffusion and capillary action. However, due to their limited terrestrial habitats, non-vascular plants have never dominated much of the Earth’s terrain.
Pre-Lab Questions
1. When did vascular plants evolve? How was this timeline uncovered?
2. How does a vascular system help a plant to grow bigger?
3. In 2 - 3 sentences, discuss how gas exchange affects transpiration.
4. Describe one way in which plants can adapt to water supply or environmental factors.
5. Why is water referred to as the “universal solvent”?
6. Why is a water droplet shaped like a sphere?
7. In essay form, discuss three adaptations plants have made to thrive in different environments.
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Experiment 1: Functionality of the Water Column
Materials
(2) 100 mL Beakers
flat on the bottom)
Permanent Marker
*2 Celery Stalks
Ranunculus root Digital Slide Images
*Cutting Board
15 - 20 Drops Red Dye
*Kitchen Knife
Ruler
*Water
Tilia Stem Digital Slide Images
*Bowl (Large enough to fit two pieces of celery
*You Must Provide
Procedure
1.
Examine the four Ranunculus root and Tilia digital slide images. In the space following the images, draw what you see. Label the xylem and phloem (refer to Figure 1 for help).
Xylem
Phloem
Figure 5: Ranunculus root (100X)
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Xylem
Phloem
Figure 6: Ranunculus root, 400X
Xylem
Phloem
Figure 7: Ranunculus root, 1000X
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Xylem
Phloem
Pith
Figure 8: Tilia (two year old stem; 40X).
Phloem
Secondary
Xylem
Primary
Xylem
Figure 9: Tilia (two year old stem; 100X).
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Phloem
Figure 10: Tilia (two year old stem; 400X).
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Include Your Drawings Here:
2. Fill a bowl with room temperature tap water.
3. Place the two celery stalks in the bowl. Use a knife to carefully cut ½ inch off the bottom of two celery stalks. This step should be performed with the celery underwater.
4. Leave one stalk submerged in the water in which it was cut, and let the other air dry for 15 minutes.
5. Use the permanent marker to label each of the 100 mL beakers as 1 or 2.
6. Fill Beaker 1 and Beaker 2 with 50 mL of water.
7. Place 8 - 10 drops of the red dye into each beaker.
8. After 15 minutes have passed, carefully, cut the tops off each stalk to create pieces which are approximately 10 cm. in length.
9. Place one stalk into each beaker with the freshly cut ends submerged in the water.
10. Observe the stalks for 12 hours, and answer the Post-Lab Questions.
Post-Lab Questions
1. How far did the dye travel in the celery which was submerged in water for 15 minutes? How far did it travel in the piece which was air-dried for 15 minutes?
2. In which celery stalk was the water column broken?
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Experiment 2: Water Movement in Flowers
Carnations colored green for St. Patrick’s Day or blue for the Fourth of July profit from transpiration to introduce color to the flowers. In this experiment, you will create your own colored carnation to gain a better understanding of how water travels through a plant.
Materials
(1) 100 mL Beaker
*Sharp Knife
8 - 10 Drops Red Dye
*Cutting Board
*You Must Provide
*1 White Carnation
Procedure
1. Fill a beaker with 50 mL water. Add 8 - 10 drops of the red dye. Under running water, cut the stem so that only one cm is left beneath the flower head. If this step is not performed under water, air bubbles can enter the xylem and prevent water from traveling to the flower.
2. Float the flower in the dye solution so that the fresh/exposed portion of the cut stem remains under water.
3. Set the beaker containing the flower in an illuminated place for 24 - 48 hours (depending on light intensity). Create a hypothesis to predict what will happen to the flower over the next two days. Record your hypothesis in the Post-Lab Questions section.
4. Observe the flower, and answer the Post-Lab Questions.
Post-Lab Questions
1. Record your hypothesis from Step 3 here. Be sure to address how the dye will affect the flower over time, and why.
2. In which part of the flower is the dye located? How did it get there?
3. What factors can influence the rate of transpiration in flowers?
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Lab 22
Plant Reproduction
Plant Reproduction
Learning Objectives
•
Compare and contrast vascular and non-vascular plant reproduction
•
Compare and contrast structure and function of bisexual and unisexual plants including gymnosperms and angiosperms
•
Identify plants as non-seed forming and seed forming
•
Explain alternation of generations during a plant’s life cycle
•
Apply double fertilization to angiosperms
Introduction
The plant kingdom encompasses many different types of plants. Embryophyta is a subkingdom of the plant kingdom which includes easily recognizable land plants such as mosses, ferns, flowering plants, and gymnosperms. Land plants have multicellular embryos which develop within the plant. They are further divided into vascular and non-vascular plants (or, bryophytes). Bryophytes encompass the oldest plant lineage, and are estimated to be approximately 475 million years old. Certain embryophytes form seeds to reproduce, while other types release spores. For example, bryophytes and some seedless vascular plants, such as fern, release spores. Vascular seed plants have adapted to a variety of habitats due in part to the robustness of the seed. Note that the common green algae is not an embryophyte. Unlike land plants which often (although not always) risk drying out, green algae is very hydrated as it absorbs water and minerals from its environment.
However, green algae can not be revived should it ever dry out, whereas many bryophtes are revivable if rehydrated within a sufficient time period.
Plant Life Cycle: Sexual Reproduction
The life cycle of all embryophytes, vascular and bryophytes, includes a diploid generation and a haploid generation. The diploid generation is called a sporophyte, and the haploid generation is called a gametophyte.
Diploid sporophytes produce hardy, haploid spores using meiosis. These spores become gametophytes by dividing and maturing. Gametophytes produce gametes by mitosis. Two gametes from different organisms come together and produce a zygote. The zygote develops into a sporophyte and the cycle continues. This cycle is the process for sexual reproduction in plants.
Alternation of Generations
The fluctuation between mitosis and meiosis is known as the alternation of generations. The time spent in each generation of the cycle varies by species within the plant kingdom. For example, the dominant generation in bryophytes is haploid. However, the dominant generation in most other plants is diploid.
There are similarities in how plants reproduce, but many differences are based on the size, form, and dispersion method of their gametophytes and sporophytes. For example, flowers, the sexual organs of angiosperms, vary greatly by size and shape, therefore requiring different methods of reproduction. Because plants
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Plant Reproduction are immobile, angiosperms (read more about angiosperms later in this lab) must utilize other, creative methods to seek out sexual partners.
Early plants including moss and ferns use water or wind to transport sperm for sexual reproduction. Further developed plants which possess formed vascular systems have adapted to other more complex methods of sexual reproduction. For example, some flowers express brightly-colored attract insects to cross-pollinate. Other flowers express caffeine molecules in their nectar. This caffeine is thought to improve memory in honeybees, which improves the flower’s likelihood Figure 1: Dandelion releasing seeds. of repeat pollinations.
Unisexual vs. Bisexual Reproduction
Plant reproduction can be unisexual or bisexual. Bisexual reproduction occurs when a plant incorporates structures which are both male and female. Unisexual reproduction occurs when a plant incorporates structures which are either male or female.
Mosses and liverworts are unisexual plants that do not form seeds to reproduce
(Figure 2). This class of plants has developed both male and female reproductive organs. The female organ is the archegonium where the archegoniophore (egg or female gametophyte) resides. The male organ is the antheridium where the antheridiophore (sperm or male gametophyte) resides. The antheridiophore depends on water for movement (dew, rain) and male and female plants must be in close proximity to each other for successful fertilization.
Gymnosperms
Gymnosperms (e.g. pine trees) are bisexual plants that produce naked seeds. Gymnosperms are called the naked seed plants
Figure 2: Moss reproduction cycle. because they are not encased by the mature ovary (fruit) as other seed-forming plants are. Although each plant contains both the male and female reproductive structures, they are in physically different locations. Typically the female reproductive structures
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(seed cones) are near the top of the tree while the male reproductive structures (pollen cones) are near the bottom of the tree. These plants are called “wind pollinated” because they rely on the wind to blow pollen grains (male gametes) to the ovule that contains the female gametes. Pollination occurs when the pollen grains come in contact with the ovule. The pollen grains germinate and pollen tubes will grow towards the egg. Once the tube reaches the egg, sperm are released from the grain and fertilize the egg. The zygote
(seed) will then mature and fall from the seed (Figure 2).
Angiosperms
Angiosperms, or flowering plants, are vascular, bisexual plants. Both the male and female reproductive organs are found in a single flower. While these plants can selffertilize, cross-fertilization (pollen from a different plant) is important to maintain genetic diversity. The carpel is a specialized organ, unique to angiosperms, that encloses the ovule and eventually matures into fruit following fertilization (Figure 3). Angiosperms are called covered seed plants because the developed seed (zygote/embryo) is within the fruit.
Angiosperms have additional distinct reproductive organs called flowers. The gametophyte is reduced to the female embryo sac, with as few as eight cells and the male gametophyte developed from the pollen grains.
Pollen grains, the male gametophyte, developed to protect the sperm during the transfer from male to female
Figure 3: Flowering plant reproductive organs. parts. Double-fertilization is also unique to these plants.
Like the gymnosperms, a pollen grain that reaches the carpel will germinate. Pollen tubes grow towards the ovary and two sperm are released. One sperm nuclei fuses with the egg to form the zygote (2n). The other nuclei unites with two haploid polar cells (in the ovary) to form a triploid (3n) cell that develops into the endosperm, a nutrient-rich tissue surrounding the seed (fruit).
Pre-Lab Questions:
1. Identify one reproductive difference between sporophytes and gametophytes.
2. How do gymnosperms vary from other, seed-forming plants?
3. How do abiotic factors such as wind or water influence pollination behavior?
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Experiment 1: Observation of Archegonium and Antheridium
Mosses are the most common group of unisex, non-vascular plants, and are common in moist environments.
Long sections of the moss Mnium containing the gametangia are useful to gain a better understanding of the moss life cycle. The antheridium is a haploid structure involved in the production and storage of sperm cells.
Alternatively, the archegonium is a structure involved in the production and storage of ova.
Materials
Mnium moss Archegonium Digital Slide Images
Mnium moss Antheridium Digital Slide Images
Procedure
1. Examine the digital slide pictures of the archegonium (Figure 4, 5, and 6). Look for the archegonium, a structure with a long neck and rounded base. Also, search for a single-celled egg within the archegonium.
Label those structures.
2. Examine the digital slide picture of the antheridium (Figures 7, 8, 9, and 10). Sperm-forming tissue will be visible within the antheridia. Label significant structures.
Egg
Archegonium
Figure 4: Mnium archegonium (100X).
Post
-Lab Questions
1. Are the spores produced by the moss sporophyte formed by meiosis or mitosis? Are they haploid or dip314
loid?
Plant Reproduction
Egg
Archegonia
Figure 5: Mnium archegonium (400X).
Egg
Archegonia
Figure 6: Mnium archegonium (400X).
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Plant Reproduction
Gametophyte
Tissue
Antheridia
Figure 7: Mnium antheridia (40X).
Antheridia
Figure 8: Mnium antheridia (100X).
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Plant Reproduction
Sterile Jacket
Antheridia
Spermatogenous Tissue
Figure 9: Mnium antheridia (400X).
Sterile Jacket
Antheridia
Sperm Forming Tissue
Figure 10: Mnium antheridia (1000X).
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Plant Reproduction
Experiment 2: Observation of a Flower
In this experiment you will observe slide images of flower reproductive parts and compare them to an actual specimen, Materials
Lilium Ovary Digital Slide Images
Lilium Anther Digital Slide Images
*You Must Provide
*Fresh Flower (Lily is recommended)
Procedure
1. Study the fresh flower. Locate and identify the important parts of the flower’s reproductive system. Sketch the flower in the space below; be sure to label the reproductive components.
2. Examine the digital slide images of the anther and ovary. Pay special attention to the labeling and review the annotations.
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Plant Reproduction
Anther sac with pollen
Figure 11: Lilium anther (40X).
Pollen Grains
Pollen Sac
Figure 12: Lilium anther (100X). Pollen grains (microspores) are clearly visible within the pollen sac (microsporangium).
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Plant Reproduction
Pollen Grains
Pollen Sac
Figure 13: Lilium anther (400X).
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Figure 14: Lilium anther (1000X).
Plant Reproduction
Pollen Grains
Ovum
Ovule
Ovary Wall
Pollen Sac
Figure 15: Lilium ovary (40X).
Embryo
Sac
Ovules
Ovary Wall
Figure 16: Lilium ovary (100X).
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Plant Reproduction
Ovule
Embryo
Sac
Figure 17: Lilium ovary 400X.
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Plant Reproduction
Post-Lab Questions
1. What are some unique characteristics of angiosperms?
2. How many carpels are in the lily? How many stamen?
3. Describe the male gametophyte of a seed plant.
4. Describe the female gametophyte of a seed plant.
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Lab 23
Invertebrates and Vertebrates
Invertebrates and Vertebrates
Learning Objectives
•
Compare and contrast the major structures and functions of invertebrates and vertebrates
•
Identify phyla as invertebrates or vertebrates: Porifera, Cnidaria, Platyhelminthes, Annelida, Arthropoda, Mollusca, Echinodermata, Chordata and Vertebrata
•
Identify structural differences and similarities between major classes of vertebrates
•
Classify organisms as displaying radial, bilateral symmetry or asymmetry
Introduction
The kingdom Animalia comprises millions of species, ranging from the snail to the hippopotamus, the ant to the elephant, the centipede to the human. Though there are notable differences in body shape and function, almost all species are motile, multicellular eukaryotes. Unlike plants, animals cannot produce energy from sunlight and, as heterotrophs, they acquire energy by consuming organic material (other plants and animals).
Symmetry
Symmetry is a useful characteristic for animal classification and represents the balanced division of an animal’s form. Radial symmetry, as is seen in starfish, is a division originating in the center and protruding outwards that produces even and balanced sections. This is similar to the divisions that are made when a pie is cut into many even pieces. Bilateral symmetry, as is seen in beetles, occurs when animal structure can be divided into two mirror images by a center line that runs through the entire object (Figure 1). No equal divisions can be detected in asymmetry, as is seen in a sponge.
Invertebrates
Animals are classified as invertebrates or vertebrates based on their internal structure. Invertebrates are organisms that lack a backbone. They make up over 98% of all animal species. Examples include the jellyfish, insects, or worms. Vertebrates pos- Figure 1: The rhino beetle exhibits bilateral sess an endoskeleton (an internal skeletal structure) and a spi- symmetry along the central axis. nal column. A backbone or spinal column is a significant adaptive advantage that enables vertebrates to occupy different ecological niches (roles). Examples include humans and dogs.
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Major Invertebrate Phyla
There are over 35 phyla in the Animal Kingdom. The vast majority of organisms fall within the following eight phyla, seven of which are invertebrates.
1. Phylum Porifera consists of sponges, considered to be the oldest of the animal phyla (Figure 2). As stationary filters, sponges play an important role in aquatic systems, feeding on particles and bacteria in the water. Porifera literally means pore bearing which accurately describes the exterior surface of the sponges.
Water enters the sponge through small pores and exits via larger openings. Nutrients are filtered out during the unidirectional flow of water through the sponge. Their asymmetric body is a loose assembly of cells (no tissues) that is supported by a matrix of collagen fibers and spicules. Sponges reproduce both sexually and asexually. 2. The phylum Cnidaria includes jellyfish, corals, sea anemones, and hydras (Figure 3). They are thought to be the first animals to develop nerves and muscles and typically alternate between two body forms: the free-swimming medusa and the stationary polyp. Both body types consist of three layers of tissue surrounded by tentacles with stinging cells containing tiny, toxic harpoons that can be used in either defense or offense.
In fact, the name “cnidarian” is derived from the Greek word for stinging nettles. They exhibit radial symmetry with a hollow body cavity to digest food. Like sponges, cnidaria can reproduce both sexually and asexually.
Figure 2: Porifera
Figure 3:Cnidaria
3. The phylum Platyhelminthes (flatworms) includes freshwater planaria, colorful marine polyclads, parasitic tapeworms and flukes. They are some of the simplest bilaterally symmetrical organisms with a defined head and tail, and a centralized nervous system containing a brain and nerves.
They lack both a body cavity and circulatory system, but do have a tubular mouth, an excretory system and a highly branched digestive system. Clusters of light-sensitive cells make up their eyespots. Recent molecular studies using DNA sequences have aided in the classification of many of these organisms which may eventually lead to a reorganization of this phylum. They are hermaphroditic, capable of both sexual and asexual reproduction.
4. The phylum Annelida is represented by marine worms (polychaetes), earthworms and leeches (Figure 4). They are bilaterally symmetric with segmented body cavities. These cavities are often represented as a tube within a tube design. Each segment has tiny hairs or bristles called setae which help the organism to move. Segmentation was an important development
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Figure 4: Annelida
Invertebrates and Vertebrates that provides distinct regions to specialize in different tasks. A one-way digestive tract, closedcirculatory system, and central nervous system also differentiate this invertebrate. Annelids play a significant ecological role by reworking soil and sediments. Annelids primarily reproduce by sexual means with some members capable of both sexual and asexual reproduction.
5. The phylum Arthropoda, the most diverse and numerous of the Ani-
mal Kingdom, includes insects, crustacean, spiders, millipedes, and centipedes (Figure 5). All arthropods have segmented bodies and are covered in a hard, flexible exoskeleton. The muscles from their jointed appendages attach to the inside of this protective cover. Many species, such as dragonflies which start as larvae and develop into winged adults, exhibit multiple life cycles. Their open body cavity contains tissues, organs and a complete digestive tract. The vast majority Figure 5: Arthropoda of members of arthropoda reproduce sexually.
6. The phylum Mollusca includes clams, snails, slugs and the octopus (Figure 6). Though there is a great deal of diversity within this phylum, all mollusks have soft bodies, many of which are covered by a hard calcified shell. The shell is produced through secretions from a layer of tissue called the mantle (also known as the pallium). A muscular foot provides locomotion and grasping. A coarse, file-like organ (the radula) allows most mollusks to drill into their prey or snag fish. Many hunt by propelling water through a siphon either for locomotion or to capture food. A mantle cavity houses gills and one-way digestive system. Ongoing molecular studies may lead to a reorganization of this phylum as more information becomes available. Most mollusks have separate sexes and reproduce sexually, although some mollusk speFigure 6: Mollusca cies are hermaphrodites.
7. The phylum Echinodermata includes sea stars, sea lilies, sea urchins, sea cucumbers, and over 6,000 other salt water species (Figure 7). Instead of bilateral or radial symmetry, most echinoderms exhibit a five part morphological symmetry. Their hard, flexible bodies are composed of small calcium plates that are often spiny and covered by a thin skin.
In fact, the name “Echinodermata” means spiny skin. Internal to the spiny skin, there is a complete digestive system and a special fluid-filled system that operates tube feet (which sometimes grow back if lost) which allow them to move, feed, and respire. Most organisms in this phylum reproduce sexually with a few specific species known to reproduce asexually. Figure 7: Echinodermata
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Invertebrates and Vertebrates
8. The phylum Chordata includes organisms that are invertebrates and organisms that are vertebrates unlike the above seven phyla which include exclusively invertebrates. Although it may seem unusual to have a phylum containing both invertebrates and vertebrates, there are four very specific embryonic characteristics or traits that allow for this classification. That is, all chordates possess each of these traits at some point during their embryonic development. First, all chordates develop a notochord. This rod-like, supportive structure extends the length of the body. Second, a dorsal nerve cord running parallel with the notochord is established. The last two traits are the development of both a muscular tail that extends beyond the anus and gill slits.
Chordates have bilateral symmetry, segmented muscles, a protective layer (feathers, scales, hair, fur, etc.,) and gill clefts (the structures located behind the mouth and in front of the esophagus).
The invertebrate chordates are the tunicates and the lancelets, both are types of marine organisms. Tunicates, more commonly known as sea squirts, look like sponges but have a notochord with a dorsal nerve cord gill slits and a tail. Lancelets are small fish-like creatures that possess the embryonic structures allowing for the classification of chordate. There are over 20 different species of lancelets.
Keep in mind that, as with many classifications, some vertebrate species might be included within the subphylum chordate in some resources, but not in other resources. For example, hagfish do not have true vertebrae but they do have a cranium. Based on the lack of true vertebrae, hagfish do not technically belong in this subphylum, but they often are classified within this group. To circumvent this issue, the term Craniata is frequently used. Craniata include animals within Chordata that have a skull.
Vertebrates
Vertebrates possess an internal skeletal structure called an endoskeleton and a spinal column. This is a key differentiating feature between vertebrates and invertebrates, which do not possess a hardened endoskeleton.
An endoskeleton is a significant adaptive advantage that enables vertebrates to occupy different ecological niches (roles) and engage in a variety of unique animal behaviors. Examples of the over 60,000 vertebrate species include humans, dogs, and the llama shown in Figure 8.
Although vertebrates and invertebrates are very different for many reasons, there are four very specific embryonic traits that all chordates possess at some point during their development. These traits include: the development of a notochord, a dorsal nerve cord, gill slits, and a muscular tail that extends beyond the anus.
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Figure 8: Llamas are historical companions to mountain trekking humans. They are often referred to as pack animals due to their ability to distribute weight over their vertebrae and back.
Invertebrates and Vertebrates
Fish and Tetrapods
The two main groups of the subphylum vertebrata include fishes and tetrapods (amphibians, reptiles, birds, and mammals). Fish are aquatic and lay soft eggs. They are cold blooded and use gills to breathe. As the name suggests, tetrapods include all animals with a four-legged body plan such as mammals and amphibians. Amphibians, such as frogs and salamanders, are found in both water and on land and lay soft eggs. Amphibians are cold blooded and breathe using gills, lungs and through their skin.
Amniotes, Reptiles, and Birds
The development of amniote eggs provided animals (amniotes) with the capability to move away from water.
Amniote eggs possess internal membranes that keep embryos moist. The two major divisions within amniotes are the reptiles (non-bird reptiles and birds) and mammals. Reptiles live almost entirely on land and lay somewhat hard shelled eggs. They are cold blooded and breathe through lungs. Birds are found on land
(and in the air) and lay hard shelled eggs. Birds are warm-blooded and breathe through lungs.
Mammals
Mammals, including humans (Homo sapiens), are normally found on land and most give live birth. They are warm-blooded and breathe through lungs. Mammals are also characterized by the presence of hair on their bodies and their ability to produce milk in modified sweat glands
(mammary glands) for their young. There are roughly 5,000 species of mammals.
There are three mammalian groups or lineages. Monotremes are mammals that lay eggs and appear more similar to reptiles with their somewhat leathery shell. Monotremes produce milk which is excreted through openings found on the mother’s skin due to the absence of nipples. The duck-billed platypus and two types of spiny anteaters are the only three monotremes species.
Marsupials are pouched mammals and include kangaroos, koalas and opossums. Marsupials give live birth to developmentally premature (or, altricial) young. The young make their way to the mother’s ventral pouch. They continue developing while being nourished by milk supplied by the mother through nipples located in the pouch.
Placental mammals develop an organ, called the placenta, during pregnancy which acts to transfer nutrients and gases between the mother and the embryo. Placental mammals include primates, all marine mammals and rodents. In fact, over half of the Figure 9: Newborn joeys are typically apover 4,000 placental mammals are rodents. proximately the size of a jelly bean.
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Invertebrates and Vertebrates
Homo sapien
Arguably the most dominant vertebrate is Homo sapien (humans). Humans are thought to be some of the longest living mammals, though other species, such as the elephant and whale are also long-lived. Though there are obvious differences between human beings and other placental mammals, there are also many similarities. Major differences include skeletal changes that have allowed for humans to stand and walk upright on two legs (bipedalism). Being bipedal also allowed for hands to become adapted to more specialized and novel roles. Human backbones are slightly S-shaped, which helps keep the body aligned over the hips. The human skull also differs so that it sits on top of the backbone with the spinal cord exiting the skull inferiorly.
Figure 10: Human spines are naturally S-shaped. However, excess curvature may result in scoliosis.
Pre-Lab Questions:
1. Identify the four major characteristics of chordates.
2. List ways in which invertebrates compensate for not having a spine.
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Invertebrates and Vertebrates
Experiment 1: Symmetry in Common Objects
Symmetry can be seen in everyday objects similar to that of the symmetry seen in organisms.
Materials
Images of Common Lab Equipment
Procedure
1. Review the objects listed below (many of these can also be found in your lab kit).
2. Indicate the type of symmetry each item displays next to the image in the Post-Lab Questions section.
Hint: It is helpful to draw lines of symmetry over the objects and observe objects from different angles.
3. For each item, explain why you chose the type of symmetry you did.
a. Petri Dish
d. Wash Bottle
b. Funnel
c. Test Tube Rack
e. Graduated Cylinder
f. Tongs
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Invertebrates and Vertebrates
Post-Lab Questions
1. Identify the type of symmetry displayed for each item. Then, indicate how you came to your conclusion:
a. Petri Dish:
b. Funnel:
c. Test Tube Rack:
d. Wash Bottle with Curved Straw:
e. Graduated Cylinder:
f.
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Tongs:
Invertebrates and Vertebrates
Experiment 2: Creating a Phylum Key
In this experiment you will take a closer look at the characteristics that describe the eight major phyla.
Materials
Table 1
Table 2
Procedure
1. Table 1 contains all of the main features of the eight major phyla discussed in the Introduction. Organize and identify which characteristics belong to which phyla in the key. The number of rows in the key represents the number of characteristics that fall into each phylum.
2. Record your results in Table 2.
Table 1: Phylum Characteristic Table
1
2
3
4
5
A
Bilateral phylum with segmentation
Most have a calcium containing shell
Five part symmetry
Specialized cell, but no tissues
Mantle of tissue covering the body
B
Hollow body cavity for food
Three tissue layers, no body cavity
Setae used for movement Jaws and skulls part of evolution
Complete digestive tract
C
Parasites
Water flows through canals of body
The first to have jointed legs
The first phylum to fly
Some have stinging cells
D
More complex because of more DNA
Body design is a tube within a tube
Tube feet
First muscle and nerves Internal skeleton
E
All live in the sea Simple animals with bilateral symmetry
Entrance and exit the same in the digestive tract
All have vertebral column
Champions of variations in appendages F
Has the most species Have spines covered with a thin skin
Some stationary polyps Some are mobile medusa Some propel using their siphon G
Muscular foot used to move
Stationary animal
Tubular mouth at the mid-body
Humans
Hard but flexible bodies with small plates
H
Spicules are the skeleton
Radula used to feed
No symmetry
Exoskeleton
Their burrowing has affected the global climate
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Invertebrates and Vertebrates
Table 2: Phylum Taxonomic Key
Porifera
Cnidaria
Platyhelminthes
Annelida
Arthropoda
Mollusca
Echinodermata
Chordata
Post-Lab Questions
1. Were any of the features used in more than one Phyla? If so, give an example of an organism from each phyla that shares the feature.
2. List three features from Table 1 that describe a Monarch butterfly (Danaus plexippus)? Using the features you listed determine what phyla it is classified in.
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Experiment 3: Taxonomy
Materials
(6) Organism Images
Procedure
1. Identify which phylum each of the following organisms belongs in. Next to each, list the criteria used in your determination.
a.
b.
c.
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Invertebrates and Vertebrates
d.
e.
f.
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Invertebrates and Vertebrates
Experiment 4: Owl Pellet Dissection
Birds of prey often consume their prey items whole, rather than selecting for the digestible components.
However, proteolytic enzymes present within the bird’s stomach are not capable of digesting all of the different structures found in an organism, and often can’t break down items such as hair, bone, teeth, bones, or exoskeletons. As a result, owls (like many other predatory bird species) produce and regurgitate pellets which contain many of these indigestible items approximately 18 - 20 hours prior to consumption. Very small bones or bone fragments occasionally pass through the pyloric sphincter and proceed through the digestive system, but pellets often contain complete animal skeletons.
In this experiment, you will probe a barn owl (Tyto alba) pellet to learn more about the skeletal system’s general physical characteristics. Note that some of the bones which you recover from the pellet will appear homologous to a human skeleton (e.g., the ribs and vertebra), while others will vary significantly (e.g., the skull). Materials
Construction Paper (Black)
*Paper Towel
Forceps
*Soap
Hand Lens
*Water
Owl Pellet
Ruler
*You Must Provide
2 Toothpicks
Caution: The owl pellets have been heat-treated but may still harbor microbes. Wear protective glasses, gloves, and apron when performing this experiment. Thoroughly wash your hands and all work surfaces with warm water and soap after complete.
Procedure
1. With your gloves on, unwrap the aluminum foil from the owl pellet and set the pellet atop the black construction paper. This will make it easier to identify the bones in the pellet as you dissect.
2. Measure the dimensions of your pellet. Record your data in Table 3.
3. Review the physical characteristics of the pellet such as texture, color, scent, etc. Be sure to use the wafting technique to detect the scent of the pellet (see the Appendix for a description of this technique).
Record your observations in Table 3.
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Invertebrates and Vertebrates
4. Carefully use the toothpicks and forceps to probe the pellet. If your pellet feels very firm, submerge it in a beaker filled with water for 1 - 3 minutes to soften it.
5. Isolate any bones you come across.
6. Continue to probe the pellet for the bones, removing fur and debris from the bones. Note that the bones will be fragile. Be careful not to break any of the bones during the dissection.
7. Use Figure 12 to identify the bones found in the pellet. Record the bones you identify in Table 3.
8. Try to recreate the skeleton of one animal found in the pellet by organizing the bones in the arrangement of the skeletal system. Pay particular attention to where the joints might have been located.
Note: Each pellet is likely to contain bones from several different animal species. Be sure to refer to
Figure 8 to complete this step.
9. Dispose of the pellet in a trash receptacle in a location which is safe from children or animals. Then, wash your work station with warm water and soap.
Table 3: Owl Pellet Data
Pellet Characteristics
Pellet Length (cm):
Pellet Width (cm):
Physical Observations:
Bone
Skull
Jaw
Scapula
Rib
Vertebrae
Hindlimb
Forelimb
Pelvic bone
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Animal Source
Number of Bones
Invertebrates and Vertebrates
Figure 12: Owl pellet bone identification chart.
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Invertebrates and Vertebrates
Post-Lab Questions
1. What did you find in your owl pellet?
2. What have you learned about the ecosystem in which the owl lives?
3. What can you infer about the nature of the community in which the owl lives?
4. How can scientists use owl pellets to study small mammals in a specific ecosystem?
5. Other birds of prey produce pellets as well, and the contents are dictated by where the bird lives. What would you expect to find in the pellet from a shorebird, such as a gull?
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Invertebrates and Vertebrates
The following lab exercise is intended to be an add-on for the Invertebrates Lab.
You will NOT have the supplies to perform the next experiment unless your teacher has specifically requested that dissection materials be included within your kit.
Experiment 5: Invertebrate Dissection
The starfish is not a fish at all, rather an invertebrate that possesses no internal skeleton. Members of the phylum Echinodermata, starfish are unique in that they are deuterostomes (instead of protostomes like earthworms, grasshoppers, clams, etc.). Deuterostomes exhibit incomplete segmentation, and a brain and spinal cord above the gut, among of other differences with protostomes. Starfish have no front or back, and can move in any direction without turning.
Note: When performing a dissection, remember these important safety notes:
•
Dissect with the scalpel or scissor blade cutting away from you (and your lab partner).
•
If you develop an allergic reaction to the preserving fluid, contact your healthcare provider. Also, inform your instructor and inform him/her of your situation.
•
Contact your local waste management company for instructions for the proper disposal of your specimen. •
Wash your hands, dissection tools, and all work surfaces with soap and water when finished with the dissection.
Materials
Dissecting Pins
Hand Lens
Dissecting Tools
Preserved Starfish
Dissecting Tray
Procedure
1. Examine the external anatomy of the starfish. The side in which the mouth is located is called the oral surface (ventral side). The opposite side is called the aboral surface (dorsal side). See Figure 13 for reference.
2. Run your gloved finger over the surface and note the texture. Use the magnifying glass to examine the spiny skin in detail.
3. Along with the stiff spines, you may also see small, hair-like gills used by the starfish to take in oxygen.
4. Pedicellaria are tiny pincers that look like pliers that are used to grip small objects.
5. Place the starfish ventral side up and note the tube feet that run down the arms, or rays, of the starfish.
They will be located on either side of the groove that runs from the tip of each ray to the center.
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Figure 13: The aboral (left) and oral (right) surfaces of the starfish.
6. Find the mouth in the center of the starfish.
7. On the oral surface of each arm are open ambulacral grooves extending from the mouth to the tip of each arm. Locate the abulacral groove running from the center down each ray.
8. Using the magnifying lens, examine the tube feet with protruding suckers on either side of the abulacral groove. 9. Flip the starfish so the dorsal side is facing up.
10. Note the eyespots at the tip of each arm, which allows the starfish to sense and respond to light. Eyespots are pigmented regions near the tip of the arms, and are covered by a transparent cuticle. To see the eyespots, spread the tube feet at the tip of the ray and examine it closely with the magnifying lens.
11. The flat central disk is at the center, and tiny, hollow, finger-like gills cover the body of the starfish.
12. The opening of the water-vascular system is called the madreporite. It is a large, button-like structure on the central disk.
13. The anus is in the center of the disk.
14. Using a scalpel, cut one inch from the tip of one of the rays. Study the cross section of the stump.
15. Note the ossicles (part of the endoskeleton) on the dorsal surface, the largest of which called the ambu-
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Invertebrates and Vertebrates lacral ossicles, which support the ambulacral groove and provide attachment for the tube feet.
16. Remove the skin from the top of this ray using dissecting scissors or scalpel. Do the same for another ray, and also cut a circular lap of skin from the central disk being careful to keep it as shallow of an incision as possible. 17. Note the feathery-looking digestive glands called the pyloric caeca. These glands make enzymes that help digest food in the stomach, located under the central disk. The thin sac lying just above the stomach is the intestine. From there, the rectal pouches store small amounts of wastes before leaving through the anus on the dorsal side of the starfish.
18. Remove the pyloric caeca from one ray, and observe the gonads underneath.
19. Remove the gonads to visualize the water vascular system. This is an internal water pressure system.
Water enters the system through the madreporite, passes through a series of canals until it reaches the tube feet. When the ampulla contracts, water is forced into the tube foot, extending it and allowing it to grab on to a surface.
20. Running the length of each ray is a lateral canal, to which tube feet are attached.
21. In the central disk, the five lateral canals connect to the ring canal.
22. Note the stone canal connecting the ring canal to the madreporite, where water enters. These canals are difficult to locate without disrupting them, but see if you can identify them.
23. With a magnifying lens, examine the inside wall of the ray to see the supporting ridges, the bulb-like ampullae, tiny sacs that create suction of the tube feet. You may also notice tiny openings in the inner wall.
These pores connect with a fill tube and are part of the external gills that help the starfish to breathe.
24. As with any biological scraps, it is best to contact your local waste management company for proper disposal procedure.
Post-Lab Questions
1. What are some common animal traits that a starfish does not possess?
2. Explain why the starfish is not classified as a vertebrate species.
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Invertebrates and Vertebrates
Your lab kit will NOT contain the following materials unless specifically requested by your instructor!!!
Experiment 6: Vertebrate Dissection
Frogs are a member of the Amphibia class of vertebrates. In many respects, the anatomy of the frog is similar to human anatomy. Thus, the study of frog anatomy is a useful tool for scientists. As amphibians, frogs may live some of their adult life on land, but return to water to reproduce.
Materials
Dissecting Pins
Flashlight
Dissection Tools
Preserved Grassfrog
Dissection Tray
Ruler
9 cm Fishing Line
Toothpick
Note: To determine the sex of your frog, examine the fingers on its foreleg (arm). A male frog typically exhibits thick pads below the thumbs.
Procedure
1. Place the frog dorsal (back) side up in the dissecting tray. Observe the external anatomy of the skin.
Don’t forget to use your sense of touch in this observation (with gloved hands of course)!
2. Locate the following features:
•
External nares (nostrils)
•
Two tympani (eardrums)
•
Two eyes, each with three lids (the third lid is a transparent covering on the eye)
3. Measure and record the length of the frog in Table 4.
4. Break a toothpick so that you have a two cm piece. Place this in the frog’s mouth to prop it open so you can observe the structures of the frog’s mouth.
5. Using the flashlight, locate the following features:
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•
Internal nares
•
Eustachian tubes
•
Opening of the esophagus
•
Glottis
•
Tongue
Invertebrates and Vertebrates
•
Two kinds of teeth; maxillary teeth help the frog to grip while vornerine teeth point inward.
6. Use forceps to grab the tongue and locate where it attaches to the floor of the mouth.
7. Insert the end of the fishing line into one of the Eustachian tubes and watch the tympanum on the dorsal side of the frog. This system allows air pressure to be equalized in the frog’s head.
8. Place the frog in the dissection tray ventral (belly) side-up, and pin the arms and legs to the tray to stabilize the specimen.
9. Using forceps and dissecting scissors, cut along the midline of the body starting at the cloaca (the urogenital opening) as shown in Figure 14. Make shallow cuts so internal organs are not damaged when cutting through the muscle and breastbone.
10. Make horizontal cuts near the arms and legs, as shown in Figure 14.
Figure 14: The dorsal (above) and ventral (below) sides of the frog. Cut lines are shown on the picture below (green).
11. Pin the flaps to the dissecting tray to expose the internal organs.
Note: If your specimen is female, you may need to remove the eggs and enlarged ovary to view the internal organs.
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12. Using a probe and forceps, lift the internal organs around so you can locate the following organs:
•
Fat Bodies: Located just inside the abdominal wall, these long, thin organs are yellow or orange in color. You may remove these structures if they obstruct your view of underlying organs.
•
Peritoneum: Directly under the body wall, this attached membrane of thin tissue forms a lining around the internal organs. There are various membranes that hold the organs in place, called mesenteries, which are also part of the peritoneum.
•
Liver: A three-lobed organ that sits high in the body cavity. It is the largest organ visible, and is dark brown in color.
•
Heart: A triangular structure that sits above the liver. Note the thin sac (called the pericardial sac) that covers this organ and the vessels extending from it. The pericardial sac can be cut to expose the heart.
•
Lungs: Two spongy organs located beneath the liver.
•
Gall Bladder: This bright green organ can be visualized by spreading apart the lobes of the liver.
•
Esophagus: A tube that transports food from the mouth to the stomach. Insert your probe into the frog’s mouth and observe where it leads.
•
Stomach: Under the left side of the liver, this bag-like digestive organ is the first site of chemical digestion in the frog.
•
Small Intestine: A long coiled tube that serves as a conduit for food and the place where nutrient absorption into the bloodstream takes place. Notice the blood vessels running through this organ.
•
Large Intestine: A widening of the small intestine signals the start of the large intestine. This may be located underneath the small intestines.
•
Spleen: Located in the middle of the body cavity, this dark red, spherical organ stores blood. You may need to lift the stomach and small intestines to see.
13. Using dissecting scissors, remove the stomach from the body cavity. Cut it open to investigate any remains from the frog’s last meal. Record observations in Table 4
14. Remove the small intestines from the body cavity. Measure the length of the small intestines and record in Table 4.
15. Continue to locate the following organs:
•
Kidneys: Dark, bean-shaped organs that filter wastes from the blood. You may have to lift the intestines to see them, as they are towards the back of the body cavity.
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Invertebrates and Vertebrates
•
Testes (for male specimens): Located above the kidneys, these round organs are typically light in color. •
Oviducts (for female specimens): Curly tubes around the kidneys.
•
Bladder: A bag-like organs that stores urine. Try to trace the tubes from the kidneys to the bladder.
16. As with any biological scraps, it is best to contact your local waste management company for proper disposal procedure.
Table 4: Grassfrog Measurements and Analysis
Frog Length
Small Intestines Length
Stomach Contents
Post-Lab Questions
1. Classify the specimen you just dissected, starting with Kingdom and ending with Species.
2. Describe the appearance of five organs you found in the frog.
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Lab 24
Animal Structure
Animal Structure
Learning Objectives
•
Identify patterns in the organization of organisms including cells, tissues, organs and organ systems
•
Explain how the major structures of the integumentary, skeletal and muscular systems influence their function
•
Compare and contrast the four major types of tissue: epithelial, connective, muscle, and nerve
Introduction
Cells are the fundamental unit of life in every organism. The trillions of cells which comprise the human body are categorized into over 200 different types. Networks of these interconnected cells unite to form tissues capable of performing specialized functions. Although there are over 200 cell types, there are only four tissue types in the human body:
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•
Epithelial tissue - Covers every surface of the body, both internally and externally. Examples include the external layer of skin, as well as the lining of the stomach.
•
Connective tissue - Connects parts of the body. Examples include ligaments and tendons.
•
Muscle tissue - Enables motion and performs specialized functions. Examples include biceps
(move your arm), cardiac muscle (pump blood) and the walls of the digestive tract (force nutrients into bloodstream).
•
Nerve tissue - Provides communication throughout the body. Examples include sensory neurons
(transmit information) and motor neurons (produce impulses to stimulate movement).
Did You Know...
Stem cells are undifferentiated cells that have not yet generated the structures or proteins characteristic of a specialized cell type. Embryonic stem cells are totipotent, meaning they have the potential to become any type of cell in the body (skin, heart, lung, bone, eye, etc.). Adult stem cells are pluripotent, meaning they are already partially specialized. For example, the stem cells in bone marrow can form blood, bone, cartilage and fat, but not a heart.
Adult stem cells are found in relatively small numbers and are thought to maintain and repair tissues. Cell-based therapies induce stem cells to differentiate and repair damaged or destroyed tissues. The goal is for these therapies to be effective in treating conditions such as corneal regeneration, Parkinson’s Disease, Alzheimer’s
Disease, multiple sclerosis, cancer, brain and spinal cord repair, and skin grafts.
Tissue types form organs, which perform specialized functions. Examples of organs in the human body include the heart, kidney, spleen, pancreas, lungs and so forth. Organ systems are comprised of multiple or-
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Animal Structure gans that collectively perform more complex functions. For example, both the bladder and kidneys are a part of the urinary system. There are eleven different organ systems in the human body. The integumentary, skeletal, and muscular systems provide structure and protection to vertebrates.
The Integumentary System
The integumentary system is comprised of skin, nails, hair and some membranes. The skin is the largest component of this system (and largest organ in the body), whose functions include:
•
Protect the body
•
Retain water
•
Host sensory receptors (temperature, pain, and pressure)
•
Maintain body temperature
Hair Shaft
Sweat Pore
Sebaceous gland Arrector pili muscle
Hair follicle
Pacinian corpuscle
Sweat gland
Figure 1: Diagram of skin.
The skin is comprised of three layers:
1. Epidermis - The outermost layer of the skin composed of thick, waterproof cells.
2. Dermis - A more complex layer below the epidermis that houses the sweat glands and hair follicles (hair roots). It has abundant connective tissue which provides elasticity and resilience.
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Animal Structure
3. Subcutaneous Layer - The inner most layer of skin, composed of fat storage cells. It protects and insulates the inner organs.
Figure 2: Skin Layers
The Skeletal System
All animals (both invertebrates and vertebrates) have a support structure that provides protection and allows for movement. Though the systems differ, their functions are similar.
There are three types of skeletal systems:
•
Hydrostatic skeletons - A primitive skeletal system found in soft bodied invertebrates, such as worms or jellyfish, and consists of a fluid filled cavity surrounded by muscle.
•
Exoskeletons - A tough and rigid outer covering, found in many insects.
•
Endoskeletons - A tough and rigid internal support structure that also serves a number of functions (Figure 3), including:
• Provide rigidity
• Provide an attachment site for muscle
• Protect internal organs
Figure 3: The endoskeleton
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Animal Structure
• Allow movement
• Produce blood cells (bone marrow)
• Store minerals (e.g. calcium)
The two primary components of the skeletal system, cartilage and bone, are often connected by tendons (a tough band of connective tissue that join muscle to bone) and ligaments (similar composition to tendons but connect bone to bone). Cartilage is stronger than tendons and ligaments, but is more flexible. It provides a low friction surface and shock absorption for joints.
The primary component of the human skeletal system is bone, which on average accounts for 15% of our total body weight. In addition to providing structure and strength to our body, the inside of bones store bone marrow, which produce a variety of cells critical to normal body function and also repair (e.g. blood cells, fat cells, blood vessels).
There are two types of bone:
•
Compact bone - The hard outer layer of all bones that appears smooth and white. Compact bone comprises about 80% of bone mass •
Spongy bone - A porous structure inside compact bone that houses red bone marrow, supplies nutrients to the bone and produces new blood cells for the body. It is part of most large bones, including the pelvis, skull and vertebrae.
Spongy
Bone
Compact
Bone
Figure 4: Types of bone.
Real World: Arthritis
Arthritis is caused by joint inflammation, and is often seen in the hands, knees, or hips. There are two primary branches of arthritis: rheumatoid arthritis, and osteoarthritis. Rheumatoid arthritis is caused be an autoimmune disorder, whereas osteoarthritis is caused by natural wear and tear experienced by joints in everyday life. Symptoms of both types include joint pain, redness, swelling, and stiffness. The cartilage which coats the ends of the bones connected by the joint, decreases over time and may even result in bone-on-bone grinding in severe cases. As to be expected, osteoarthritic symptoms typically worsen as the body ages. In rheumatoid arthritis, the decrease in cartilage may be accelerated by
Figure 5: Common areas for arthritis. local attacks from the immune system on the lining of the joint capsule.
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Animal Structure
Muscular System:
Muscles enable motion and perform vital functions (e.g., digestion). The approximately 630 muscles in the human body fall into three types and account for 40% of total body mass (on average).
•
Skeletal Muscle: Also known as striated muscle, range considerably in size and shape, have alternating dark and light bands and are connected to bones, enabling motion. Unlike other types of muscle, it is voluntarily controlled. For example, if you want to jump, these muscles contract so you can. Figure 6: Innervation of skeletal muscle (left) and muscle fiber anatomy (right).
•
Smooth Muscle: Found in almost all hollow cavities in the body (e.g., stomach, intestine, bladder, uterus), are smaller than skeletal muscle, do not exhibit striations and are involuntarily controlled
(they function without conscious thought). For example, digestion, which is facilitated by smooth muscle, occurs whether you think about it or not.
•
Cardiac Muscle: Found only in the heart, has alternating striations and is involuntarily controlled.
Cardiac muscle never rests until the heart stops beating.
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Animal Structure
Table 1: Major Systems Matched with Function and Component
System
Function
Components
Nervous
System
Receives and processes information and then directs the body of what to do
Brain, nerves, spinal cord, sensory organs
Endocrine
System
Works with the nervous system, using chemicals (hormones) to communicate with the body and regulate its chemical processes Many glands (pituitary, thyroid, adrenal, etc.), pancreas
Lymphatic
System
Defense system of the body as well as providing transportation of many fluids and compounds
Spleen, lymph nodes, lymphatic vessels, tonsils, thymus
Muscular
System
Moves the body
Skeletal muscle, smooth muscle, cardiac muscle
Skeletal
System
Provides protection and support for the body Bones, ligaments, cartilage, tendons
Digestive
System
Processing and absorption of nutrients from food
Salivary glands, liver, stomach, gallbladder, esophagus, pancreas, teeth, small and large intestines, rectum
Circulatory
System
Transport oxygen, nutrients and other compounds throughout the body and then takes away waste products from the body
Heart, blood, blood vessels
Respiratory
System
Obtains oxygen and transfers waste gases out of the body
Lungs, nose, trachea, diaphragm, pharynx, larynx, bronchi
Urinary System
Excretion of waste (urine) from the body
Kidney, bladder, ureters, urethra
Integumentary
System
Covers and protects the body
Nails, skin, hair, some membranes
Reproductive
System
Prepare cells for and participate in reproduction
Males: penis, testes, seminal vesicles, prostate gland, epididymis, vas deferens
Females: vagina, uterus, fallopian tubes, ovaries 358
Animal Structure
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Did You Know...
Aerobic activity results in a lot more than calorie burn-off! In fact, as soon as you start exercising a biological domino effect takes place. Here’s the breakdown:
1. Your body begins breaking ATP down into ADP and phosphate ions. When these bonds break, energy is released which can be used for muscle contraction.
2. The initial store of ATP is used up within a minute or two. To harness more, glycogen, glucose, and fat molecules are broken down as an alternative energy source to create ATP.
3. Breaking down these molecules requires extra oxygen. To achieve this, heart rate increases and blood is pulled away from unneeded systems like the digestive system. This allows the body to pump more blood quickly, pushing red blood cells throughout the body which drop off oxygen molecules to the muscles.
4. Every time an ATP, glycogen, or glucose molecule is broken down energy is released. Some of this energy is used by the muscles to accommodate the exercise movements, but the energy also results in an increase in body temperature.
5. To maintain a healthy body temperature, the circulatory system comes into play again by focusing blood-flow towards the skin.
This may cause redness, and releases heat as sweat glands are activated to release heat-laden moisture.
6. When a workout is complete, the brain releases a rush of endorphins which contributes to the “runners high” many people feel.
Pre-Lab Questions:
1. Give two examples of how the integumentary system, skeletal system and muscular system interact. 2. Hypothesize how the muscular system works with the circulatory system to maintain homeostasis.
3. Relate tissues to organs and organ systems. Give an example.
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Animal Structure
Experiment 1: Examining Skin, Bones and Muscle Histology
The skin (the primary component of the integumentary system) is a large organ which provides protection to the body. Muscles and bones are located beneath the skin and enable movement, structure, and further body protection. In this experiment you will observe the structure and function of cells of the integumentary, skeletal, and muscular systems.
Materials
Skin Digital Slide Images
Cardiac Muscle Digital Slide Image
Cortical (Compact) Bone Digital Slide Image
Skeletal Muscle Digital Slide Image
Trabecular (Spongy) Bone Digital Slide Image
Smooth Muscle Digital Slide Image
Note: All images are located at the end of the procedure.
Procedure
Part 1: The Skin
1. Observe the digital slide picture of the hairy animal at 100X, 400X, and 1000X.
2. Note the locations of the epidermis, dermis, dermal papillae, and the sweat glands. Note that fat cells that comprise the subcutaneous layer.
Part 2: Investigating Cortical (Compact) vs. Trabecular (Spongy) Bone
1. Examine the digital slide images of the cortical (also referred to as compact) and trabecular (also referred to as spongy).
2. Be sure to read through and locate all of the labeled features.
Part 3: Muscle Structure
1. Examine the digital slide images of cardiac, skeletal, and smooth muscle.
2. Again, review all of the labels and annotations presented with each image.
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Animal Structure
Human Skin 100X.
Epidermis
Layer
Sweat gland duct
Dermal Papillae
Dermis Layer
Human Skin 400X. The stratum corneum contains primarily keratinized cells. Dermal papillae can form ridges that aid in gripping (fingerprints).
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Animal Structure
Human Skin - A 1000X. The layers of the epidermis can be distinguished. The keratinocytes of the stratum granulosum contain dark staining granules.
Hair follicle
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Human Skin- B 1000X. Enlargement of a hair follicle located in the dermis. The keratinized hair root and the hair matrix (region of cell division) are enclosed by the hair follicle.
Animal Structure
Haversian system Haversian canals
Interstitial lamellae
Concentric lamellae Cortical Bone (Compact bone) 100X. Cylindrical units called osteons are notable in cortical bone.
Lacunae containing osteocytes
Canaliculi
Lamella
Cortical Bone 1000X. Osteocytes, mature bone cells, are located in cavities called lacunae. Canaliculi provide passageways for nutrients traveling to the osteocytes.
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Animal Structure
Trabeculae
Ma
r
ro w Ca v it y
Trabecular Bone (Spongy Bone) 100X. Marrow fills the internal cavities found in trabecular bone.
Trabeculae
Marrow Cavity
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Trabecular Bone 1000X.
Animal Structure
Branched structure
Cardiac Muscle – A Tissue 100X. Note the branched structure.
Cardia
c mus cle cell nu cleus Inte r cala ted di
scs
Cardiac Muscle Tissue 1000X.
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Animal Structure
Cardiac Muscle Tissue – B 100X. Cardiac muscle is well known for its location and striations. It is only found as a component of the heart walls. They are branched in structure and possess one nucleus (as opposed to skeletal muscle which is cylindrical in shape and multi-nucleated). Cardiac muscle also possess intercalated discs which hold the cardiac muscle fibers together when the heart undergoes contraction.
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Animal Structure
Muscle Fibers
Skeletal Muscle 100X. The multinucleated skeletal muscle fibers run parallel to each other.
Nuclei
Single Muscle
Fiber
Skeletal Muscle 1000X. Striations resulting from the arrangement of contractile proteins that run perpendicular to the muscle fiber.
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Animal Structure
Smooth Muscle Tissue 100X. Smooth muscle tissue surrounds the gastrointestinal tract and helps. Similarly, it also helps blood vessels contract and push blood through the circulatory system.
Smooth Muscle Tissue 1000X. These cells can be described as spindle-shaped.
Note the elongated ends of the cells.
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Animal Structure
Post-Lab Questions
1. Label the arrows in the following slide image:
A
B
C
Human Skin 400X.
2. Which layer comprises the majority of skin?
D
Explain how you came to your conclusion using your
knowledge of skin components and functions.
3. What component of the subcutaneous layer provides protection to the underlying organs?
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Animal Structure
4. What layer of skin causes dandruff? Is this the same layer that peels off after a sunburn? How do you know? 5. Wrinkles occur with age due to a loss of elasticity and collagen. Which layer of skin do you believe is most responsible? Do you think topical ointments that claim to firm skin really work? Explain why or why not? 6. Sketch a drawing of a bone (you pick!), and label the compact and spongy regions.
7. Identify at least two differences and two similarities between cortical and trabecular bone. These may be anatomical or physiological.
8. Why do you think spongy bone allows more flexibility than compact bone?
9. Suppose you are looking at an unlabeled slide image of muscle. List two items which would help you determine if the muscle was cardiac, smooth or skeletal?
10. Why does muscle account for such a large portion of your body mass? Explain using scientific evidence.
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Animal Structure
Experiment 2: Calcium in Bones
As a child, you may have been encouraged to drink milk so that “your bones would grow strong.” Milk is high in calcium, one of the primary components of bones. On average, 99% of all of your body’s calcium is stored in bones. If your body becomes calcium deficient, it will use the reserve stored within your bones.
Ultimately, your bones can lose their strength if this reserve is depleted.
Remarkably, bones are fairly light weight but can resist large amounts of stress. In addition to calcium, collagen fibers provide strength and flexibility, enabling bones to withstand physical pressure, resistance, impact, tec. If these components are damaged, the structure of the bone may also change.
In the following experiment you will examine the importance of calcium in bones. Egg shells possess calcium, and acetic acid (vinegar) removes calcium from bone.
Materials
Figure 3: Endoskeleton
200 mL Acetic Acid (Vinegar), C2H4O2
Permanent Marker
Aluminum Foil
*Water
(2) 250 mL Beakers
Construction Paper (Black)
*You Must Provide
*1 Egg
Note: This lab requires five days of observation. Wear gloves when handling egg shells.
Procedure
1. Use the permanent marker to label the two 250 mL beakers as 1 and 2.
2. Measure and pour 200 mL of water into Beaker 1.
3. Measure and pour 200 mL of acetic acid into Beaker 2.
4. Carefully crack one egg into two, approximately equal sizes. Discard the yolk and gently rinse the egg shells under running water.
5. Use a gloved hand to gently place the first half of the egg shell in Beaker 1. If possible, try to submerge the egg shell by filling any vacant cavities with water. Cover the beaker with aluminum foil and let sit at room temperature for five days.
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Animal Structure
6. Use a gloved hand to gently place the second half of the egg shell in Beaker 2. Again, try to submerge the egg shell. Cover the beaker with aluminum foil let sit at room temperature for five days.
7. Divide the black construction paper in half. Label one side as water and one side as acetic acid.
8. On the 5th day, remove the acetic acid and water from Beaker 1 and Beaker 2, respectively.
9. Use a gloved hand to remove each of the egg shells from the beakers, and place them one the corresponding side of the construction paper.
10. Visually examine the physical characteristics of the egg shells. Record your observations in Table 2.
11. Use your fingers to gently apply pressure to the egg shells. Notice the differences in the strength or flexibility. Record your observations in Table 2.
Table 2: Effect of Acetic Acid on Egg Shells
Beaker
Observations
Beaker 1: Water
Beaker 2: Acetic
Acid
Post-Lab Questions
1. What do these results suggest about the role of calcium in bones? Is it important? Why or why not?
2. Why did the flexibility of the egg shell change after it was removed from the acetic acid?
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Animal Structure
Experiment 3: Agonists and Antagonists
Many muscles work in antagonistic pairs; meaning, when one muscle contracts, another relaxes. For example, when the bicep contracts it lifts the forearm up and lengthens and at the same time. The forearm is forced down and the bicep lengthens (causing it to relax) when the tricep contracts.
There are two primary types of muscle contractions: isotonic and isometric. Isotonic contractions occur when the length of a muscle changes. Isometric contractions do not change the muscle length.
Materials
Figure 3: Endoskeleton
Figure 7 (located at end of procedure)
Procedure
Part 1
1. Identify and mark as many antagonistic pairs as possible for the following body regions (hint, it will help to feel the muscles on your own body to determine the antagonistic pairs):
A. Lower leg
B. Upper leg
C. Lower arm
D. Upper arm
Record your pairings in Table 3.
2. Lay your left arm on a flat surface with the palm up. Place your right palm on top of your left bicep and slowly raise your left forearm. Notice the shortening of the bicep muscle. That is an isotonic contraction.
3. Stand, hold your left arm in front of you and slowly squeeze your fist. Notice that the muscles in the forearm do not shorten when contracted. That is an isometric contraction.
Part 2
4. For the following exercises, perform the muscle contraction and determine if it is an isotonic or isometric contraction. a. Place your hand underneath the table. Push up on the table up while keeping your arm straight.
b. Lay on your back on the floor. Pull your chest up to your knees (a sit up).
c. Sit in a chair and place a ball (or other object) between your feet. Slowly lift your feet into the air.
d. Lie on your side on the floor and raise your upper leg towards the ceiling.
e. Sit in a non-moveable chair and place your feet straight out in front of you against a solid object
(like your desk). Try to push away the desk.
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Animal Structure
Figure 7: Human Muscle
Table 3: Antagonistic Muscle Pairings
Body Region
Lower Leg
Upper Leg
Lower Arm
Upper Arm
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Paired Muscles
Lab 25
The Circulatory and Respiratory Systems
The Circulatory and Respiratory Systems
Learning Objectives
•
Explain the function of the heart and blood in the circulatory system
•
Describe the structure and function of the respiratory system highlighting respiration
•
Explain how the circulatory and respiratory systems work together to maintain homeostasis
Introduction
All cells and ultimately systems, in the human body require nutrients and oxygen. They must also be capable of eliminating accumulated waste products. The circulatory and respiratory systems play a major role in these processes.
The Heart
The circulatory system is driven by the heart. As the heart contracts, blood is propelled from the left ventricle into the aorta. From there, it travels through successively smaller arteries before reaching arterioles, capillary beds, and finally tissues. Together with the heart, the circulatory system is responsible for distributing blood and nutrients to all the cells of the body.
Blood
Figure 1: The heart is divided into two chambers containing four regions: right atrium, right ventricle, left atrium, and left ventricle.
Blood, a fluid tissue, is a mixture of plasma, red blood cells, white blood cells, and platelets (Figure 2). The blood that circulates throughout the veins and arteries of the body is made of a variety of cells and cell fragments suspended in a liquid called plasma. Plasma makes up the largest portion of blood, accounting for 55% of the fluid. Plasma itself is nearly 90% water. Table 1 lists different types of cells found in blood and their corresponding functions.
Blood serves three main functions:
1. Transport oxygen, carbon dioxide, nutrients, hormones, and waste.
2. Regulate pH, body temperature, and water content of cells.
Figure 2: Red and white blood cells attacking a pathogen.
3. Protect against blood loss (through clotting) and disease through phagocytic white blood cells and antibodies.
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The Circulatory and Respiratory Systems
Table 1: Types of Blood Cells
Erythrocytes
Also known as red blood cells; comprise approximately 45% of blood. Contain hemoglobin for oxygen and carbon dioxide transport.
Lymphocytes
Involved in the immune response; produce antibodies.
Monocytes
Thrombocytes
Phagocytotic macrophages.
Also known as platelets. Involved in blood clotting.
Neutrophils
Key to wound healing.
Eosinophils
Phagocytosis.
Basophils
Leukocytes
Release histamines.
Also known as white blood cells; comprise approximately 1% of blood. Involved in fighting disease and foreign objects in the body as part of the immune system.
Red blood cells have special regions that bind and transport respiratory gases (O2 and CO2). This exchange of gases and nutrients is known as respiration. White blood cells are one component of the body’s defense system that attack foreign organisms or particulates including bacteria and viruses. They also produce antibodies. Platelets are involved responsible for clotting blood.
The Circulatory System
The circulatory system is a closed system where the blood remains in the vessels (arteries and veins) and does not flow freely in the organs. Blood flows throughout two circuits, both of which provide a mechanism for gas exchange: the pulmonary and the systemic (Figure 3).
The pulmonary system adds oxygen (O2) to the blood and removes carbon dioxide (CO2). Alveoli are small nodules located on deep braches of the lung that have a large surface to volume ratio. The gas exchange takes place between these nodules and the hemoglobin, a protein in red blood cells. The systemic system delivers oxygenated blood to the cells through capillaries and removes waste products, including CO2. All cells reside within close proximity to capillaries to facilitate this process. Both systems are driven by the heart.
378
Figure 3: The circulatory system showing the pulmonary system in red and the systemic system in blue.
The Circulatory and Respiratory Systems
Oxygen depleted blood travels through the vena cava into the right atrium. The right atrium contracts, forcing the blood into the right ventricle. The right ventricle contracts, pushing the blood into the pulmonary vein where it travels to the lungs and exchanges CO2 for O2. The blood then returns to the left ventricle.
The ventricle contracts, ejecting the blood into the left atrium. The left atrium contracts and pumps blood into the aorta, which delivers it to the rest of the body. A pulse is created when blood is pumped by the heart into the aorta (systolic pressure). Blood pressure on the vessels (when heart muscle relaxes) creates the diastolic pressure. These two measurements are taken as your blood pressure in a doctor’s office to monitor the health of your circulatory system.
Figure 4: Anatomy of the heart.
379
The Circulatory and Respiratory Systems
The blood also makes its way to the digestive system where it picks up nutrients and to the urinary system, where it disposes of waste products. Together, the respiratory system and circulatory system provide the body with oxygen, and a delivery route for nutrients.
Cells require oxygen to produce ATP by cellular respiration. The process produces CO2 as a byproduct, a toxin that must be removed. The respiratory process relies on the diffusion of gases (O2 in and CO2 out) across the cell membranes (Figure 4).
The Respiratory System
The main organ of the respiratory system is the lung. This spongy, elastic tissue stretches and contracts as you breathe. Air travels through your nose or mouth to the trachea, which branches into many smaller tubes called bronchi and then bronchioles. This airway leads to microscopic air sacs called alveoli, which are covered with capillaries. This is the first site of gas exchange.
Respiration involves three steps to transport the oxygen (with the help of the cardiovascular system) and carbon dioxide throughout the body.
Pulmonary ventilation is the movement of air between the atmosphere and the lungs. It occurs during inhalation and exhalation. The diaphragm and the muscles of the thoracic wall change the space and pressure inside the lungs. When the diaphragm contracts, it leaves more space for the lungs to expand and lowers the internal air pressure. Air pressure is greater outside of the body, and will move to a region of lower pressure (inside the lungs). External respiration is the movement of oxygen from the alveoli into pulmonary capillaries and carbon dioxide from the pulmonary capillaries to the alveoli. Oxygen and carbon dioxide are transported between the lungs and body tissues. Internal respiration is the movement of oxygen from the capillaries into body cells, and of carbon dioxide from the body cells into ca- Figure 5: The trachea subdivides into pillaries. bronchi, bronchioles, and smaller
The steps involved in respiration are:
380
•
Ventilation
•
Gas exchange between alveolar air and lung capillaries
•
Bulk flow of the circulation
•
Gas exchange between the capillaries and tissue cells
•
Cellular consumption and production of gases
conduits that conduct air from the external environment to the alveoli, the site where gas exchange occurs.
The Circulatory and Respiratory Systems
Molecules of oxygen and carbon dioxide are passively exchanged through the process of diffusion into and out of the blood. Diffusion is the net movement of molecules from a region of higher concentration to a region of lower concentration as a result of their random movement.
Figure 6: Gas exchange: oxygen (red) diffuses from the lungs to the blood stream and is used in aerobic respiration. Carbon dioxide (green and yellow), a waste product of respiration, diffuses from the bloodstream to the lungs where it is exhaled out of the body.
Pre-Lab Questions
1. Identify one reason why carbon dioxide is considered a waste product to the body.
2. How does pressure in the lungs change when a person inhales? How does it change when a person exhales?
3. Explain the role of hemoglobin in the circulatory system in 1-2 sentences.
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The Circulatory and Respiratory Systems
Experiment 1: Understanding Lung Mechanics
In this experiment, you will model lung function.
Materials
Balloon
Wash Bottle
Rubber Band
Procedure
1. Attach the balloon to the end of the straw that fits inside the wash bottle, and secure with a rubber band.
The balloon will act as the lung.
2. While gently squeezing the bottle, replace the top and secure tightly.
3. Squeeze the bottle and record what happens to the balloon in Table 2.
4. Release the bottle and observe what happens in Table 1.
Table 1: Understanding Lung Mechanics Observations
Squeezed Bottle Observations (Step 3)
Released Bottle Observations (Step 4)
Post-Lab Questions
1. What happens to the lung (balloon)? Why?
2. What would happen if the seal at the base of the bottle leaked?
3. What causes a collapsed lung?
4. Is a collapsed lung functional? Why or why not?
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The Circulatory and Respiratory Systems
Experiment 2: Observation of Blood
Materials
Digital Slide Images of Blood
Procedure
1. Examine the digital slide picture of blood. Then answer the Post-Lab Questions.
Blood Smear 100X
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The Circulatory and Respiratory Systems
Blood smear 1000X. Blood cell types shown include erythrocytes (red blood cells), lymphocytes (the sparse, large, pale pink circles), and neutrophils.
Typical red blood cells
Sickled red blood cells
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Red Blood Cells 1000X. Red blood cells (RBCs) are typically circular in shape. Patients with sickle cell anemia produce blood cells with atypical cell shapes; these tend to appear elongated or suckered near the middle of the cell.
The Circulatory and Respiratory Systems
Post-Lab Questions
1. What is the most prevalent cell type in blood? Hypothesize why that is.
2. Based on what you have learned, how would you explain carbon monoxide poisoning?
Experiment 3: Modeling the Circulatory System
In this experiment you will explore and model the structures and function of the circulatory system.
Materials
2 Check Valves
60 cm of Tubing
2 Drops of Dye
*Scissors
Nail
*Water
Permanent Marker
2 Plastic Bottles
*You Must Provide
Procedure
1. Use the permanent marker to label the plastic bottles 1 and 2.
2. Use a nail to carefully puncture two holes in the side of each bottle (approximately 3.5 cm from the bottom of the bottle) so that the holes are across from each other and at the same height from the bottom of the bottle. It is best to puncture the bottle with the nail straight through both sides. This will help keep the holes level.
LAB SAFETY: BE EXTREMELY CAREFUL WHEN PUNCTURING THE BOTTLE. KEEP IN MIND
WHERE YOUR HANDS ARE AT ALL TIMES.
3. You may need to maneuver the nail around the hole to make it large enough to fit the plastic tubing
(Figure 7). The goal is for the tubing to fit tightly in the hole.
4. Carefully cut the 60 cm tubing into four equal lengths of 15 cm.
5. Attach one end of one piece of 15 cm tubing to the OUT end of a check valve.
6. Attach one end of a second piece of tubing to the IN end of the check valve.
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The Circulatory and Respiratory Systems
Figure 7: Maneuver the nail in a circular motion to enlarge the hole so that the tubing fits tightly.
7. Repeat Steps 3 and 4 for the remaining check valve.
8. Construct a closed system by inserting the free ends of the tubes into the holes you created in the plastic bottles to create a loop. The tubes should be far enough into the bottle so that water can flow from tube to bottle to tube. The tubes in each bottle should not touch (Figure 8).
Note: Properly assembled models should have the direction of the check valves facing in opposite directions.
9. Fill each bottle ¾ with water. The water level should be higher than Figure 8: Model Set Up the tubing.
10. Place the model on an underpad or paper towel to protect surfaces for the next step. This will also help catch water that leaks from the model. There should be minimal leaking if tubes are tightly in the bottles.
11. Place one drop of dye in each bottle and close the bottle by screwing on the caps.
12. Gently swirl the bottles to allow dye and water to mix. Be careful not to squeeze the bottles!
13. Note where the water is initially before squeezing the bottles and record your observations in Table 2.
14. Squeeze bottle 1 with both hands. Observe the direction of water flow and the water level in each bottle while you are squeezing and right after you stop squeezing. Record your observations in Table 2.
15. Repeat Step 13 for the bottle 2 and record your observations.
16. Squeeze one bottle and then the other back and forth. Observe the direction of water flow and the water level in each bottle. Record your observation in Table 2.
17. Draw a diagram of your model under Table 2.
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The Circulatory and Respiratory Systems
18. Use the terms Left Side of Heart, Right Side of Heart, Lungs and Body and knowledge gained from the
Introduction to label your drawing.
19. Use arrows to label your drawing to show the direction of water flow.
Table 2: Circulatory System Model Observations
Action
Water Level and Direction of Water Flow
Before Squeezing the Bottles
Squeezing Bottle 1
Directly After Squeezing Bottle 1
Squeezing Bottle 2
Directly After Squeezing Bottle 2
Squeezing Bottle 1 and 2 Back and Forth
Diagram of Model
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Post-Lab Questions
1. What did each of the bottles represent? What does the water represent?
2. What circulatory system structure do the check valves represent? What is their function in the circulatory system?
3. What do you predict would have happened if there were no valves present in the model? How would this affect the human body?
4. Where on the model did you assign the Lung and Body labels? Use circulatory system structures when answering.
5. Why use two bottles to represent the heart instead of just one? Use scientific evidence to support your answer. 6. Humans have a four-chambered heart. How could you modify the model to show this? What supplies would you need? Draw a diagram of a possible model.
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Lab 26
The Sensory and Nervous Systems
The Sensory and Nervous Systems
Learning Objectives
•
Identify the structures and function of neurons including dendrites, axons and synapses
•
Identify the major structures and functions of the central and peripheral systems
•
Distinguish between the somatic and autonomic systems of the peripheral nervous system
Introduction
The nervous system carries a large responsibility for maintaining the homeostasis within the body. It is the master control and communication center of the body. The nervous system allows the body to interpret, integrate, and react to the surrounding environment as well as the internal stimuli occurring simultaneously throughout the body. Voluntary actions such as writing, reading, or running, as well as involuntary actions such as the heart pumping blood through the body, the pancreases releasing enzymes, or the gut digesting food are monitored by this complex system.
The three main responsibilities of the nervous system are sensory input (for stimuli inside and external to the body), integration of sensory input, and a motor response by activating effector organs. To accomplish these substantial tasks, the nervous system is organized into two distinct sub-systems: the central nervous system (CNS) and the peripheral nervous system (PNS).
Special cells called neurons transmit signals to and from the brain, while surrounding glial cells maintain the neurons so they operate at their peak performance. Each neuron has a long arm called the axon, which sends signals to other neurons’ dendrites
(Figure 2). What are commonly called ‘nerves’ are really the axonal processes of nerve cells. These processes are insulated with myelin to ensure signals are sent quickly.
It is important to remember that many neurons throughout the body work together to transmit signals from the brain to tissues, and from tissues back to the brain. The junction of two neurons is called a synapse (synapse), and is the site of biochemical signal
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Figure 1: Nervous system anatomy.
The Sensory and Nervous Systems
(neurotransmitter) exchange. Many drugs target synapses and the molecules that exchange between synapses to regulate behavior.
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Did You Know...
There are as many as
1,000,000,000,000,000 synapThere are two parts to the nervous system: the central nervous system and ses in the human brain! Thethe peripheral nervous system. The central nervous system includes the se provide the power to probrain and spinal cord, and is responsible for most of the information pro- cess all of the information our brain receives from the cessing, storage, and retrieval. nerves all over the body.
The peripheral nervous system consists of all the neurons outside of the brain and spinal cord, and can be further classified by function.
•
The somatic nervous system coordinates the body’s movements and for processing external stimuli. This division of the nervous system is regulated by conscious control, and is used for activities such as running, swimming, brushing your teeth, etc.
•
The autonomic nervous system is responsible for the unconscious control of the body, and is further separated into three divisions:
The sympathetic nervous system responds to impending danger or stress, and is implicated in the “fight or flight” response. It is also responsible for control of certain activities that people never think about having to do, such as breathing, blood pressure, and heart rate.
The parasympathetic nervous system is responsible for changes observed during relaxation, such as the slowing of the heart beat, relaxation of pupils, and stimulation of digestion.
The enteric nervous system is responsible solely for digestion. This includes every action from the time food enters the mouth until it exits the body.
Motor neurons transmit messages from the spinal cord to muscles. They project beyond the spinal cord and stimulate muscles by innervating them. Sensory nerve cells transduce information about physical or chemical stimuli to the nervous system for interpretation by the brain.
The Five Senses
The five senses recognized by humans are:
1
Sight
2
Sound
3
Smell
4
Touch
5
Taste.
Figure 2: Neuron.
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The Sensory and Nervous Systems
When these senses are experienced, sensory cells signal the neurons, which transmit the information to the brain via motor neurons. The brain processes the signal and directs the effectors, such as muscles and glands. Some of these responses are voluntary, while others, such as reflexes, are involuntary.
Sensory receptors not only perceive the type of stimulus (temperature, presDid You Know... sure, sight, taste, sound), but also the intensity, location, and duration. Different receptors receive different types of stimuli and patterns that provide addi- Multiple sclerosis is a disease tional information to the brain. in which the myelin coating
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Another interesting aspect of the sensory system is vision. What is sight at the cellular level? A ray of light hits the object being viewed, then reflects into the eye through the lens. Then the light (the image) hits the retina, which has receptors that signal to the brain what color the ray is. Then, the brain receives this information and assembles the colored rays of light into an image.
on neurons deteriorates.
Without the myelin covering, the bioelectric messages sent through the neurons don’t travel far enough to transmit messages to other cells.
The retina has two kinds of receptors – rods and cones. Rods receive black and white rays, while cones detect colors. There are three kinds of cones: red, blue, and yellow. All the colors that are seen are made up of combinations from these three colors. A phenomenon called after image occurs when selected rods or cones tire out and stop working momentarily. When this occurs, the color seen is the a combination of what colors the remaining rods and cones see (usually if you stare at a blue object you will see yellow, if you stare at red you will see green, and vice versa).
4
1 = Aqueous Humor
2
3
2 =Vitreous Humor
1
3 = Sclera
4 = Choroid
10 7 6
9
4
5 = Ciliary Body
6 = Iris
7= Pupil
8
5
8 = Retina
9 = Optic Nerve
10 = Lens
Figure 3: Eye anatomy.
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The Sensory and Nervous Systems
Signals from the brain instruct the rest of the body on how to adapt to stimuli. For instance, in order to see objects at different distances, muscles in the eye contract and relax in order to change the shape of the eye to focus. When looking at a distant object, the lens in the eye is relatively flat. However, the lens becomes more round when looking at a closer object.
The brain receives large amounts of information from our surroundings via sensory organs. Sensory stimuli are detected, processed, and translated into another action through the nervous and sensory systems. The body is an amazing machine, with intricate internal systems that allow us to perceive our environment in multiple ways, and helps us to react to it accordingly in learned or automatic (genetically programmed) responses.
Pre-Lab Questions
1. Explain how the nervous system works with each of the other human body systems.
2. Hypothesize why some human body functions like breathing and heartbeat are an unconscious, involuntary effort.
3. Relate dendrites to axons and synapses.
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Experiment 1: Observation of the Nervous System
In this experiment you observe the structures of the neuron and hypothesize how this determines its function.
Materials
Nervous System Digital Slide Images
Procedure
1. Examine the digital slide pictures of neurons. Be sure to review the labels and annotations accompanying each image.
Axon
Cell Bodies
Spinal Cord Smear 100X. Neuroglial cells are closely associated with neurons as they act as supporting cells. 396
The Sensory and Nervous Systems
Neuroglial
Cells
Dendrites
Cell Body
Spinal Cord Smear 1000X.
Nerve fiber
Perineurium
Myelinated Nerve Fiber 100X.
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The Sensory and Nervous Systems
Myelinated Nerve Fiber 1000X. Schwann cells create a myelin sheath by wrapping around the fiber in a ‘jelly- roll’ fashion.
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Post-Lab Questions
1. Label the arrows for A and B in the slime image, based on your observations from the experiment:
A
B
2. Hypothesize what the neuroglial cells function is. Support your hypothesis using scientific evidence.
3. Provide a hypothesis explaining how the axon and the dendrites affect the function of a neuron.
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The Sensory and Nervous Systems
Experiment 2: Limits and Capabilities of the Eye
In this experiment you will explore the visual component of the sensory system.
Materials
Image of Blue Triangle (located at the end of the procedure) Image of Yellow Triangle (located at the end of the procedure) Ruler
*Pencil/Pen
*White Construction or Printer Paper
* You Must Provide
Procedure
Part 1
1. Hold the pencil by the eraser with the point facing the ceiling. It should be at arm’s length in front of your right eye.
2. Close your left eye, and slowly move the pencil toward your eye.
3. Focus on the end of the pencil.
4. Keep moving the pencil until the tip is out of focus. Measure the distance (called the near point) between your eye and the pencil in cm. Record your measurement in Table 1.
5. Repeat Steps 2 - 4, this time closing your right eye. Record your measurement in Table 1.
6. If you wear glasses, repeat these steps for each eye with glasses off. Record your measurements for the right and left eyes in Table 1.
Part 2
7. Quickly review the yellow and blue triangles (located at the end of this procedure).
8. Formulate a hypothesis stating what you think will happen if you stare at the yellow triangle for 30 seconds. Then, create a second hypothesis stating what you think will happen if you stare at the blue triangle for 30 seconds. Record your hypothesis in Post-Lab Question 1.
9. Stare intently at the yellow triangle for 30 seconds. Quickly shift your gaze to a sheet of white paper.
10. Repeat Step 9 using the blue triangle.
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The Sensory and Nervous Systems
Table 1: Left and Right Eye Measurements
Eye
Near Point (cm)
Near Point (without glasses, if applicable; cm)
Right Eye
Left Eye
Post-Lab Questions
1. Include your hypotheses from Part 2: Step 7.
2. Examine Table 2. According to the table, how “old” are your eyes?
Table 2: Age Point Correlation Table
Age (years)
10
20
30
40
50
Near Point (cm)
9
10
13
18
50
3. Identify the two primary fluids found within the eye. Which one of these fluids provides nutrients to the rest of the eye? Hypothesize why this is important.
4. What did you see after you stared at the yellow triangle, and then looked at the white paper? What happened when you looked at the blue triangle?
5. Use scientific reasoning to explain what is happening when you stare at the image, then transfer your gaze to a blank sheet of paper.
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Experiment 3: Skin Response
In this experiment you will explore how the nervous system interacts with the integumentary system.
Materials
(3) 250 mL Beakers
*Pencil/Pen
16 Paperclips
*Participant (not the student)
Ruler
*Tape
Toothpick
*Warm Water
*Ice Water
*Room Temperature Water
* You Must Provide
Note: You will need a laboratory partner to perform Steps 1 - 4 of this experiment.
Procedure
Part 1
1. Make a caliper by unwinding two of the paperclips until they are straightened. Then, bend each paperclip in the middle to form a U-shape. There should be approximately 0.5 cm between the two tips of each paperclip after being bent into a Ushape.
2. Tightly wrap each tip with a small piece of adhesive tape. This
Figure 4: Paperclip shape reference.
minimizes any sharp points on the paperclip. See Figure 4.
3. Repeat Steps 1 - 2 for the remaining paperclips with the ends set 1.0, 1.5, 2.0, 2.5, 3.0, 3.5 and 4.0 cm apart. When finished, you should have two calipers for each other the eight possible sizes.
4. To test the sensitivity of different body regions, ask your partner to close his/her eyes. Start with the largest caliper (the one with 4.0 cm between the tips) and place both tips of the paperclip onto the left side of your partner’s scalp. Do NOT tell your partner if you place one or both of the tips on his/her head.
5. Ask your partner how many points s/he can distinguish. If s/he can feel two points, move on to the 3.5 cm caliper and test again (your partner should keep his/her eyes closed!). Repeat until your partner can only feel one point. Record the caliper size at which only one point can be felt in Table 3.
6. Continue to test different regions on the left side of the body. Use Table 3 to select body regions.
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7. Repeat Steps 4 - 5 on the right side. Observe if there is a difference between the left and right sides of the body, and record your results in Table 3.
Part 2
8. Obtain three beakers. Fill beaker one with ice water, beaker two with room temperature water, and beaker three with warm water. Immerse your left pointer finger in the ice water, and your right pointer finger in the warm water for 30 seconds.
9. Then, place the finger that was in the ice water into the beaker filled with room temperature water. Note the sensation (did it feel cooler or warmer?) in Table 4.
10. Next, place the finger that was in the warm water into the room temperature water. Again, note the sensation in Table 4.
Table 3: Two-Point Discrimination Test Results
Body Region
Scalp
Forehead
Lips
Front of Neck
Back of Neck
Shoulder
Upper Arm
Elbow
Forearm
Wrist
Back of Hand
Palm of Hand
Tip of Thumb
Tip of Second Finger
Tip of Third Finger
Tip of Fourth Finger
Tip of Fifth Finger
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Left Side Caliper Measurement
Right Side Caliper Measurement
The Sensory and Nervous Systems
Table 4: Temperature Sensation
Initial Beaker/Final Beaker
Sensation
Ice Water/Room Temperature Water
Room Temperature Water/Ice Water
Post-Lab Questions
1. Which of these areas (forearm, back of the neck, index finger, and the back of the hand) contains the most touch receptors? Why is this useful?
2. What was the sensation for each hand when you placed them in the room temperature water?
Experiment 4: Brain Mapping
The brain is the control center of the body. Different areas of this organ are responsible for different functions. In the following exercise, you will learn the anatomy of the brain and the functional areas important to our everyday actions.
Materials
Figure 6
Permanent Marker
**White Swim Cap
**If you have a latex allergy, wear vinyl gloves when drawing on the cap, and do not put it on your head! Note: Feel free to use as many additional marker colors as you like.
Procedure
1. Study Figures 5, 6, and 7 noting the major functional areas of the brain.
2. Lay the swim cap on a flat surface, or you may choose to ask a volunteer to put it on while you map!
3. On the left side of the swim cap, outline the following structures:
Cerebral hemispheres (left and right)
Cerebellum
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Central sulcus (divides parietal and frontal lobes)
Lateral sulcus (divides the front and parietal lobes)
4. Outline and label the following lobes:
Frontal
Temporal
Parietal
Occipital
5. Number the functional areas of the cortex located in the lobes, using the number key on Figures 7 and 9
(lateral and superior views only).
6. Wear your cap and explain to someone what each of the functional areas you labeled does!
10
1 = Frontal Lobe
11
2 = Parietal Lobe
2
5
1
3 = Occipital Lobe
12
4 = Temporal Lobe
6
8 9
5 = Brocca’s Area
6 = Tertia Optica Area
7
7 = Wernicke’s Area
3
4
8 = Pars Triangularis
9 = Pars Opercularis
14
13
10 = Gyrus Praemotoricus
11 = Gyrus Praecentralis
Figure 5: Brain anatomy (lateral view).
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Figure 6: Sagittal plane.
Broca’s Area
Touch
Corpus Callosum
Writing
(right-handed)
Auditory Cortex
(right-ear)
Auditory Cortex (leftear)
Spatial Visualization and
Analysis
Wernicke’s Area
(language,
math, etc.)
Visual Cortex
(right-field)
Visual Cortex (left
-field)
Figure 7: Superior view. The right and left cerebral hemispheres are connected by the corpus
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callosum, allowing them to coordinate activity.
The Sensory and Nervous Systems
Post-Lab Questions
1. Describe the function of three areas of the brain.
2. Hypothesize why major functions of the human body are regulated by specialized parts of the brain.
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Lab 27
Ecology of Organisms
Ecology of Organisms
Learning Objectives
•
Define ecology as interactions amongst organisms and their environment
•
Analyze the effects of abiotic and biotic factors in an ecosystem
•
Assess how generalists and specialists define habit tolerance
Introduction
Ecology is the scientific study of both the interactions among organisms and the interactions between organisms and their environment. It is an interdisciplinary field that blends Earth science, evolutionary biology and general biology. Ecology explores the adaptations of organisms, the distribution of organisms and biodiversity.
Group of communi es
Group of popula ons
Group of conspecific organisms
Individual
Ecology can be divided into different levels or categories. Ecoform of life system ecology looks at the “big picture”. It focuses on the cycling of chemicals and the energy flow within an ecosystem. EcoTissues or systems system ecology includes all the abiotic factors and species within an ecosystem. Community ecology is concerned with the interSmallest actions between species and how those interactions affect the unit of life structure and organization of the community. As the name implies, population ecology is concerned with populations. A popu- Figure 1: Note the relationship between lation is a group of individuals of the same species living in the species, populations, communities and ecosystems. same area. Population ecology focuses on factors that affect population density and growth. Ecology of organisms, or organismal ecology, is the study of individual organisms and the adaptations that have enabled them to survive in their abiotic environments.
Abiotic and Biotic Factors
Environments encompass abiotic and biotic components. Abiotic components include the physical and chemical aspects. Physical factors that affect an ecosystem include sunlight, temperature, water currents, wind, nature of soil, latitude and altitude. Chemical factors include available nutrients (in soil or water), water salinity, toxicity concentrations, and water levels in soil. Biotic components refer to the living factors in an environment and include the producers, consumers and decomposers. Producers are the organisms that convert inorganic material to organic substances. Plants are an example of a producer. By converting sunlight and carbon dioxide to carbohydrates, they are producing food for the consumers. Consumers and decomposers are completely dependent on producers for nourishment. Since plants are an important producer, it follows that sunlight powers nearly all ecosystems (aquatic and terrestrial).
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Ecology of Organisms
Organisms have adapted and evolved anatomical, physiological, and behavioral characteristics that compensate for variation within the environment. The blending of ecology and evolutionary biology is probably most evident when examining organismal ecology. In this way, the environment shapes the organisms.
Habitat Tolerance
Adaptations that organisms have developed allow them to compensate for minimal temporal and spatial variation withFigure 2: Creatures have learned to thrive in a variety of extreme environments, such as the in their environment. This is accomplished, for example, by regulating body temperature or controlling the rate at which desert climate pictured above. water is transpired. Nevertheless, there are limits to an organism’s ability to compensate for environmental factors. No single species can tolerate all of Earth’s environments. The geographic distribution of a species is thus limited by the physical environment. That is, species distribution is limited by an environment’s abiotic factors.
Although living organisms are found all over the planet, all species have a defined habitat tolerance assigning a viable range of environmental conditions. For example, some plant species can tolerate a broad range of soil variability, while others are confined to a single soil type. If a species has a narrow habitat tolerance because of one or more abiotic factors, then they are limited in their distribution range. Species with a broad range of tolerance are usually distributed widely, whereas those with a narrow range have more restricted distribution. Species can be categorized as specialists or generalists. Specialist species are those that have a limited diet or cannot tolerate a wide range of environmental conditions. Consider a cactus. A cactus has a limited tolerance for soil conditions, water levels and temperatures. Generalists, on the other hand, have a wider range of tolerance for environmental conditions and diet.
For example, omnivores are typically generalists, whereas herbivores tend to be specialists. Interestingly, invasive species are frequently generalists.
Figure 3: Dandelion (Taraxicum officionale)- species like the dandelion
Habitat tolerance along with a species’ geographic range (limited vs. are very common and show no aspects of rarity making them very widespread) and its local population size (large vs. small) determines a common handling a broad range of species’ commonness or rarity. These classifications can be very signifi- tolerances.
cant to industries such as agricultural production and wildlife management.
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Ecology of Organisms
As mentioned above, the environment shapes the organisms. What is the effect of organisms on the environment? Environmental changes are influenced by the organisms that inhabit that environment. In this way, organisms shape the environment. A change in species distribution can modify interactions within the environment. The loss of a native species or the invasion of a non-native species can alter the survival of other organisms within the environment. Therefore, control of invasive species and conservation of endangered species are important to maintain the balance of the entire system.
Pre-Lab Questions
1. Would you expect endangered species to be more frequently generalists or specialists? Explain your answer.
2. How does temperature affect water availability in an ecosystem?
3. Choose a species and describe some adaptations that species developed that allow them to survive in their native habitat.
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Ecology of Organisms
Experiment 1: Effects of pH on Radish Seed Germination
Natural soil pH depends on the parent rock material from which it was formed and processes like climate.
Soil pH is a measure of the acidity or alkalinity of the soil. Acidic soils are considered to have a 5.0 or lower pH value whereas 10.0 or above is considered a strong basic or alkaline soil. The pH of soil affects the solubility of nutrients in soil water and thus it affects the amount of nutrients available for plant uptake. Different nutrients are available under differing pH conditions.
In this experiment we will look at the effect of pH on the germination and growth rate of radish seeds in order to determine the range of pH tolerance for the seed. Acidic or basic water will be used in order to stimulate acidity or alkalinity in soil.
Materials
2 mL 4.5% Acetic Acid (Vinegar), C2H4O2
Soda) Solution, NaHCO3
Permanent Marker
*Paper Towel Sheets (cut to fit into the petri dish)
(3) 5 cm Petri Dishes
*Scissors
3 pH Test Strips
*Sunny Location
Radish Seed Packet
*Water
Ruler
2 mL 15% Saturated Sodium Bicarbonate (Baking *You Must Provide
Procedure
1. Use the permanent marker to label the top of each of the three petri dishes as Acetic Acid, Sodium Bicarbonate, or Water.
2. Carefully cut three small circles from the paper towel sheets. The circles should comfortably fit within the bottom of the petri dish.
3. Place the circles in the dishes, and wet them with approximately 2 mL of each respective solution (acetic acid, sodium bicarbonate, or water).
4. Gently press the reaction pad of three, pH test strips onto the wet paper towels. Record your data in Table 1.
5. Arrange 10 radish seeds on each paper towel in each petri dish. Make sure the seeds have space and are not touching. Then, place the top of the petri dish on the bottom.
6. Create a hypothesis regarding which environment will provide the most hearty radish seed growth. Be sure to indicate why you believe this will be true, and use scientific reasoning. Record your hypothesis in
Post-Lab Question 1.
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Ecology of Organisms
7. Place the petri dishes in a sunny or well-lit, warm place. Be sure to keep the paper towels moist for the length of the experiment with the appropriate solution if any of the towels dry out.
8. Observe the seeds daily for seven days, and record the number of seeds that germinate in Table 1. Note when the seeds crack and roots or shoots emerge). On the seventh day, record the lengths of radish seed sprouts (mm or cm).
Table 1: pH and Radish Seed Germination
Stage/Day Observations
Acetic Acid
Sodium Bicarbonate
Water
Initial pH
1
2
3
4
5
6
7
Post-Lab Questions
1. Record your hypothesis from Step 6 here:
2. Compare and construct a line graph based on the data from Table 1 in the space below. Place the day on the x axis, and the number of seeds germinated on the y axis. Be sure to include a title, label the x and y axes, and provide a legend describing which line corresponds to each plate (e.g., blue = acetic acid, green
= sodium bicarbonate, etc…).
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3. Was there any noticeable effect on the germination rate of the radish seeds as a result of the pH? Compare and contrast the growth rate for the control with the alkaline and acidic solutions.
4. According to your results would you say that the radish has a broad pH tolerance? Why or why not? Use your data to support your answer.
5. Knowing that acid rain has a pH of 2-3 would you conclude that crop species with a narrow soil pH range are in trouble? Is acid rain a problem for plant species and crops?
6. Research and briefly describe a real world example about how acid rain affect plants. Be sure to demonstrate how pH contributes to the outcome, and proposed solutions (if any). Descriptions should be approximately 2 - 3 paragraphs. Include at least three citations (use APA formatting).
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Lab 28
Ecological Interactions
Ecological Interactions
Learning Objectives
•
Hypothesize how habitat influences niches
•
Explain how substances accumulate in trophic levels through biomagnification
•
Analyze the effects of changes in a food web
Introduction
An ecosystem is composed of communities of living organisms at a particular time. This includes plants and animals, as well as non-living elements such as water, wind, heat, chemical constituents, etc. Each organism within a community has a habitat and a niche. An organism’s habitat is the location in which it physically resides or is adapted to reside. An organism’s niche refers its function or “occupation”. Occupation defines the way in which the organism obtains and sustains all of the elements needed for survival.
Figure 1: Mountain gorillas have a restricted geographic range, a narThe dynamics of an ecosystem and the complexity of its habitats are a row habitat tolerance, and a small result of energy flow, nutrient cycling, and water. Organisms can be prolocal population. It is highly vulneraducers or consumers in terms of energy flow. These distinctions com- ble to extinction.
Trophic Levels
prise the basis for trophic levels. Autotrophs are primary producers; they make their own food typically using water, energy from the environment (e.g., sunlight) and carbon dioxide as their carbon source. In other words, autotrophs do not rely on other organisms for survival; they are self-nourishing. Autotrophs are the critical link between solar radiation or inorganic chemicals and every other planetary consumer. Autotrophs produce all the organic compounds (carbohydrates, fats, proteins) that are critically needed by all other organisms.
Phototrophs use sunlight for their energy source while lithotrophs use inorganic molecules (hydrogen sulfide, sulfur, etc.) for their energy source. Photosynthesis, for example, is used by phototrophs to produce carbohydrates from inorganic molecules as shown in the following equation:
CO2 + H2O + light → CH2O + O2
Heterotrophs are consumers and obtain energy and carbon from organic molecules made by primary producers. Heterotrophs, therefore, rely on other organisms for their nourishment. Heterotrophs are sorted into levels based on what their diet includes. These levels are: primary, secondary and tertiary. Herbivores are primary consumers whereas carnivores and omnivores are secondary consumers. Tertiary consumers are on
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Ecological Interactions top of the food chain and feed on secondary and primary consumers. They are sometime known as apex predators and can be carnivores or omnivores.
Organisms that share the same basic foods are said to be in the same trophic level (feeding level). Beginning with the autotrophs, energy “flows” through the system along a circuit called a food chain. This occurs in the form of carbon-carbon bonds. Carbon-carbon bonds are broken for energy consumption when respiration occurs. For example, corn (a phototroph) produces carbohydrates (starch) which provide humans Figure 2: Heterotrophic species consuming a phototrophic species.
(heterotrophs) with a source of energy and carbon. Consider another scenario in which an herbivore ingests the corn and a predator consumes the herbivore. The herbivore receives its nourishment from the corn and the predator receives its nourishment from the herbivore.
Biomagnification
Biomagnification is the accumulation of a substance as it works its way up the food chain by transfer of the substance from lower trophic level organisms to higher trophic organisms. Biomagnification results in higher or magnified substance concentrations for organisms higher in the food chain. Only about 10% of food calories make it from one trophic level to the next in biotic feeding systems. Energy transfer is in no way 100% efficient. Inefficient energy transfer is what accounts for the classically depicted ecological pyramid, with a wide base representing the producers and a narrow top representing the tertiary consumers.
Figure 3: Inefficient vs. Efficient transfer pyramid. There is a big difference in efficiency between direct and indirect consumption. Calories are lost as they move up the food chain, which also contributes to the fact that there are more prey than predators present in a given ecosystem.
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Ecological Interactions
Organisms at lower trophic levels eat small amounts, but higher trophic levels organisms eat many lower trophic level organisms; thus, chemicals accumulate in higher quantities. Biomagnification becomes a concern when the magnified chemicals are harmful to life or the environment. For example, mercury is not degraded or excreted by organisms. It can be absorbed by autotrophs and ingested by consumers accumulating in tissues. At each level, the concentration of mercury increases.
As mentioned above, all life on Earth is directly or indirectly reliant on primary production. Algae are extremely prevalent autotrophs that undergo photosynthesis (mainly algae) utilize aquatic carbon dioxide and water to produce carbohydrates (CH2O). Recall that photosynthesis produces carbohydrates and oxygen.
Predation and Competition
In an ecosystem , populations interact with each other in a variety of ways. The two most common interactions observed are predation and competition. Competition occurs when two or more populations need the same resource: water, food, or space. Competition can occur directly or indirectly. Predation is an interaction in which one population (prey) is a food source for another (predator).
Symbiotic Relationships
Perhaps some of the most interesting animal behaviors are the long term interactions or relationships observed in nature between two or more different species. Relationships between species range from those which favor only one of the species, to those in which both species benefit. Parasitism is an example of a relationship in which one species benefits and the other species suffers. Mutualism, on the other hand, is a relationship in which members of all involved species benefit. The term symbiosis describes the long-term relationships between organisms of different species. Keep in mind that some resources use the term symbiosis to describe only mutualistic relationships while others use it for all long-term interspecies interactions.
Pre-Lab Questions:
1. Explain how some heterotrophs could be described as either a primary consumer or a secondary consumer.
2. Where do you think fungi fit into a food chain? Are they autotrophs or heterotrophs? Explain your answer using scientific reasoning and data.
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Ecological Interactions
Experiment 1: Ecological Interactions
Materials
(1) 10 mL Graduated Cylinder
contain 1000 mL)
(1) 100 mL Graduated Cylinder
10 mL Vegetable Oil
*You Must Provide
*Large Pitcher, Jug, or Glass (must be able to
Procedure
1. Measure out 990 mL of water into your big jug using the 100 mL graduated cylinder. The jug represents the primary producers.
2. Measure 10 mL of oil and add it to the jug with 990 mL.
3. Clean the 10 mL graduated cylinder so there is no oil residue.
4. Let the oil coalesce at the top of the container, the oil is the accumulation agent. It is insoluble because we want it to represent the organism’s inability to break it down.
5. Calculate the volume of water and the volume of oil in your 1000 mL container. Calculate percent concentration of oil. Record in Table 1.
Hint: V (of two or more substances) = (V (of single substance)/ total V of the mixture) x 100
6. Pour 100 mL of the mixture into the 100 mL graduated cylinder and let the mixture settle.
7. Calculate the volume of water and the volume of oil in your 100 mL container. Calculate the percent concentration of oil. Record in Table 1.
8. Now, pour 10 mL of the mixture into the 10 mL graduated cylinder.
9. Calculate the volume of water and the volume of oil in the 10 mL graduated cylinder. Calculate the percent concentration of oil.
Table 1 Volume and Concentration Totals
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Trophic Level
Cylinder
Volume of
H2O
Volume of Oil Total Volume
1st
1000 mL
1000 mL
2nd
100 mL
100 mL
3rd
10 mL
10 mL
Percent Oil
Ecological Interactions
Post-Lab Questions
1. What is the percent concentration of oil in the first, second, and third trophic levels in our food chain?
2. How did the concentration of oil change from one trophic level to the next?
3. How does the change in concentration represent biomagnification?
4. How does this also illustrate the amount of energy transferred from one trophic level to the next?
If you would like to read more about issues surrounding bioaccumulation simply search for case studies that address POP (persistent organic pollutants) and biomagnification in ecosystems.
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Ecological Interactions
Experiment 2: Oil Spills and Aquatic Animals
Understanding ecological interactions extends beyond reviewing the natural interactions between the environmental and animal factors. In fact, many would argue that human interactions with animals can cause an equally great (or even greater) impact on animals or the environment due to the number of species which become endangered or extinct as a result of human behavior. This may transpire through hunting, habitat encroachment, pollution, etc. In this experiment, you will examine the affect of oil and detergent pollutions animal coats.
Materials
(5) 250 mL Beakers
Sponge
3 Chamois Cloth Pieces
Wooden Stir Stick
3 Feathers
*Stopwatch
(1) 100 mL Graduated Cylinder
*Large Bowl
2
2 /3 T. (apx. 40 mL) Liquid Dishwashing Soap
*Paper Towel
Measuring Spoon
*Water
2
2 /3 T. (apx. 40 mL) Vegetable Oil
Permanent Marker
*You Must Provide
2
2 /3 T. Powdered Dishsoap
2 2/3 T. Powdered Laundry Detergent
Procedure
1. Observe the texture of the chamois cloth. Try to use three to four different adjectives to describe the cloth, and record your observations in Table 3.
2. Repeat Step 1 using the feathers instead of the chamois cloth.
3. Use the permanent marker to label the 250 mL beakers as Liquid Dishwashing Detergent, Powdered
Dishsoap, Powdered Laundry Detergent, Water, Oil + Water.
4. Use the measuring spoon to add 2 2/3 T. (approximately 40 mL) amount of liquid dishwashing detergent, 2
2
/3 T. powdered dishsoap, and 2 2/3 T. powdered laundry into their respective beakers. Set the beakers
aside.
Note: Three total measurements in Step 4. One measurement should be allocated into one of the three, labeled beakers.
5. Use the 100 mL graduated cylinder to add 40 mL water to the Water beaker.
6. Add 100 mL water to the Liquid Dishwashing Detergent, Powdered Dishsoap, and Powdered Laundry Detergent beakers. Use the wooden stir stick to gently stir the mixture, being careful not to introduce unnecessary bubbles to the solutions.
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Ecological Interactions
7. Fill the large bowl approximately 50% with room temperature water. This will be your rinse bowl.
8. Add 40 mL oil and 10 mL water to the Water + Oil beaker.
9. Add the three pieces of chamois cloth to the Water beaker. Allow the cloth to soak for one minute. Note any changes to the cloth texture. Record your observations in Table 3.
10. Add the water-saturated pieces to the Water + Oil beaker. Allow it to soak for an additional three minutes. 11. Rinse the pieces in the Rinse beaker and place them on paper towels to air dry. Note any initial changes to the cloth in Table 3.
12. Repeat Steps 9 - 11 with the feathers.
13. Place one piece of chamois cloth and one feather in the liquid dishwashing detergent beaker. Allow to soak for approximately five minutes.
14. Repeat Step 11 placing items into the Powdered Laundry Detergent and the Powdered Dishwashing
Detergent beakers.
15. Use the sponge to gently wash the items in their respective beakers. Then, individually transfer the items to the rinse bowl and continue to rinse the items.
16. Place each item on a paper towel, and allow to air dry.
17. Examine the dried items and observe their final textural status. How do they compare to the initial texture? Record your observations in Table 3.
Table 3: Cloth and Feather Observations
Item Type
Initial Observations
Water-Saturated
Observations
Oil-Soaked
Observations
Final Observations
Chamois
Cloth
Feather
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Ecological Interactions
Post-Lab Questions
1. Which solution cleaned and restored the items most effectively?
2. Research the ingredients included in powdered laundry detergent, powdered dishwashing soap, and liquid dishwashing soap. What difference(s) do you notice, and how do you think this affected the soaps’ ability to restore the items?
3. Identify one marine species with an outer layer which is similar to the chamois cloth, and one which is similar to the feathers.
4. How might aquatic species be affected in a natural environment if a coat or outer shell was compromised by pollution? How might bird species be affected?
5. Identify one additional way that aquatic species may contact water pollution (other than via coat or shell contamination). 6. Aquatic life may be directly affected when oil leaks or spills occur. For example, the Deepwater Horizon oil spill in 2010 (which is considered to be the largest oil spill in petroleum industry history) affected over
8,000 marine species including fish, birds, mammals, and more. Research one species (plant or animal) that can be affected by oil spills.
426
Appendix
Good Lab Techniques
Good Lab Techniques
Good Laboratory Practices
Science labs, whether at universities or in your home, are places of adventure and discovery. One of the first things scientists learn is how exciting experiments can be. However, they must also realize science can be dangerous without some instruction on good laboratory practices.
•
Read the protocol thoroughly before starting any new experiment. You should be familiar with the action required every step of the way.
•
Keep all work spaces free from clutter and dirty dishes.
•
Read the labels on all chemicals, and note the chemical safety rating on each container. Read all Material Safety Data Sheets
(MSDS) prior to each experiment. These are provided on www.eScienceLabs.com. An underpad will prevent any spilled liquids from contaminating the surface you work on.
•
Thoroughly rinse labware (test tubes, beakers, etc.) between experiments. To do so, wash with a soap and hot water solution using a bottle brush to scrub. Rinse completely at least four times. Let air dry.
•
Use a new pipette for each chemical dispensed.
•
Wipe up any chemical spills immediately. Check MSDSs for special handling instructions (provided on www.eScienceLabs.com).
A
B
C
Special measuring tools in make experimentation easier and more accurate in the lab. A shows a beaker, B graduated cylinders, and C test tubes in a test tube rack.
429
Good Lab Techniques
•
Use test tube caps or stoppers to cover test tubes when shaking or mixing – not your finger!
•
When preparing a solution, refer to a protocol for any specific instructions on preparation. Weigh out the desired amount of chemicals, and transfer to a beaker or graduated cylinder.
Add LESS than the required amount of water. Swirl or stir to dissolve the chemical (you can also pour the solution back and forth between two test tubes), and once dissolved, transfer to a graduated cylinder and add the required amount of liquid to achieve the final volume.
•
A molar (M) solution is one in which one liter (L) of solution contains the number of grams equal to its molecular weight. Ex:
1 M = 110 g CaCl ÷ 1 L
Disposable pipettes aid in accurate measuring of small volumes of liquids. It is important to use a new pipet for each chemical to avoid contamination. (The formula weight of CaCl is 110 g/mol)
•
A percent solution can be prepared by percentage of weight of chemical to 100 mL of solvent (w/
v) , or volume of chemical in 100ml of solvent (v/v).
Ex:
20 g NaCl ÷ 80 mL H2O = 20% w/v NaCl solution
•
Concentrated solutions, such as 10X, or ten times the normal strength, are diluted such that the final concentration of the solution is 1X.
To make a 100 mL solution of 1X TBE from a 10X solution:
10 mL 10X TBE ÷ 90 mL water = 100 mL 1X TBE
•
Always read the MSDS before disposing of a chemical to insure it does not require extra measures. (provided on www.eScienceLabs.com)
•
Don’t pour unused chemical back into the original bottle.
•
Avoid prolonged exposure of chemicals to direct sunlight and extreme temperatures.
•
.Immediately secure the lid of a chemical after use.
•
Prepare a dilution using the following equation: c1v1 = c2v2
Where c1 is the concentration of the original solution, v1 is the volume of the original solution, and c2 and v2 are the corresponding concentration and volume of the final solution. Since you know c1, c2, and v2, you solve or v1 to figure out how much of the original solution is needed to make a certain volume of a diluted concentration.
430
Good Lab Techniques
•
If you are ever required to smell a chemical, always waft a gas toward you, as shown in the image above. This means to wave your hand over the chemical towards you. Never directly smell a chemical. Never smell a gas that is toxic or otherwise dangerous.
•
Use only the chemicals needed for the activity.
•
Keep lids closed when a chemical is not being used.
•
When diluting an acid, always pour the acid into the water. Never pour water into an acid.
•
Never return excess chemical back to the original bottle. This can contaminate the chemical supply.
•
Be careful not to interchange lids between different chemical bottles.
•
When pouring a chemical, always hold the lid of the chemical bottle between your fingers. Never lay the lid down on a surface. This can contaminate the chemical supply.
•
When using knives or blades, always cut away from yourself.
•
Wash your hands after each experiment.
431
Credits
Credits
Text/Photography
Chiras, D., (2010). Environmental science. (8th ed.). Sudbury, Massachusetts: Jones and Bartlett Publishers,
LLC.
Cornely, K., Pratt, C. W., (2004). Essential biochemistry. Hoboken, New Jersey: John Wiley & Sons, Inc.
Damjanov, I., McKinnell, R. G., Parchment, R. E., Perantoni, A. O., & Pierce, G. B. (2006).Biological basis of cancer. (2nd ed.). New York, New York: Cambridge University Press.
Evers, C. A., Starr, C., Starr, L., (2010). Biology today and tomorrow. (3rd ed.). Belmont, California: Brooks/
Cole, Cengage Learning.
Lewis, R., (2007). Human genetics concepts and applications. (7th ed.). New York, New York: The McGraw
Hill Companies, Inc.
Johnson, G. B., Raven, P. H., (1989). Biology. (2nd ed.). St. Louis, Missouri: Times Mirror/Mosby College
Publishing.
Marieb, E. N. (2009). Essentials of human anatomy and physiology. (9th ed.). San Francisco, California:
Pearson Education Inc.
Office of Transportation and Air Quality. (2013, 08 01).Renewable and alternative fuels. Retrieved from http:// www.epa.gov/otaq/fuels/alternative-renewablefuels/ Saferstein, R., (2013). Forensic science from the crime scene to the crime lab. (2nd ed.). San Francisco, California: Pearson Education Inc.
Tiedje, J. M., Thompson, D. K., Xu, Y., & Zhou, J. (1993). J. Zhou (Ed.), Biodiversity Prospecting Hoboken,
New Jersey: Wiley-Liss.
US Department of Energy. (2013, 09 25). Clean energy. Retrieved from http://www.epa.gov/cleanenergy/ energy-and-you/affect/coal.html 433
Student Manual
Copyright Information
Information
Copyright
Introductory Biology Lab Manual
© 2013 eScience Labs, LLC. All rights reserved. This material may not be reproduced, displayed, modified, or distributed, in whole or in part, without the express prior written permission of eScience Labs. Appropriate citation(s) must accompany all excerpts and/or quotations.
For written permissions, please contact info@eScienceLabs.com
Note, educational institutions and customers who have purchased a complete lab kit may reproduce the manual as a print copy for academic use provided that all copies include the following statement: “© 2013 eScience Labs, LLC. All rights reserved.”.
This manual was typeset in 11 Arial and 12 Chalet-London 1960. Arial font provided by Microsoft Office Suite, 2010. Chalet-London 1960 font licensed from House Industries, 2011.
The experiments included within this lab manual are suitable for supervised or unsupervised learning environments. eScience Labs assumes full liability for the safety and techniques employed within this manual provided that all users adhere to the safety guidelines outlined in the mandatory eScience Labs Safety Video, Preface, and Appendix. All users must understand and agree to the eScience Labs safety guidelines prior to beginning their lab experiments. eScience Labs does not condone use of the lab materials provided in its lab kits for any use outside of the curriculum expressly outlined within the lab manual.
3
Table of Contents
Introduction
Lab 1
Introduction to Science
Lab 2
General Lab Safety
Lab 3
Chemical Bonding Fundamentals
Lab 4
Introduction to the Microscope
Biological Processes
Lab 5
The Chemistry of Life
Lab 6
Diffusion
Lab 7
Osmosis
Lab 8
Enzymes
Lab 9
Cellular Respiration
Cellular Fundamentals
Lab 10 Cell Structure and Function
Lab 11 Mitosis
Lab 12 Meiosis
Lab 13 DNA and RNA
Lab 14 Mendelian Genetics
Lab 15 Population Genetics
Kingdoms of Life
Lab 16 Taxonomy
Lab 17 Bacteria and Archaea
Lab 18 Protista
Lab 19 Fungi
5
Table of Contents
Plant Kingdom
Lab 20 Energy and Photosynthesis
Lab 21 Plant Circulation
Lab 22 Plant Reproduction
Animal Kingdom
Lab 23 Invertebrates and Vertebrates
Lab 24 Animal Structure
Lab 25 The Circulatory and Respiratory Systems
Lab 26 The Sensory and Nervous Systems
Environmental Biology
Lab 27 Ecology of Organisms
Lab 28 Ecological Interactions
Appendix: Good Lab Techniques
6
Time and Materials
If you are allergic to nitrile, please contact eScience Labs at info@esciencelabs.com and we will send you an alternative type of safety gloves. Swim-caps and rubber bands include latex. Always wear your nitrile safety gloves when working with these items!
Please note that the times listed are approximations and may differ. Please read through the procedure and plan accordingly.
Lab 1 Introduction to Science
Time Required: 1 hour
Additional Materials: None
Lab 2 General Lab Safety
Time Required: 1 hour
Additional Materials: Water
Lab 3 Chemical Bonding Fundamentals
Time Required: 1 hour
Additional Materials: Water, Newspaper, Notebook Paper, Scissors
Lab 4 Introduction to the Microscope
Time Required: 1 hour
Additional Materials: Computer, Internet Access
Lab 5 The Chemistry of Life
Time Required: 2 hours
Additional Materials: Egg White, Hot Pad, Kitchen Knife, Microwave, Paper Towels, Pot (Small), Scissors, Stovetop, Water (Hot and Tap),
7
Time and Materials
Lab 6 Diffusion
Time Required: 2 hours
Additional Materials: Scissors, Stopwatch, Tape, Water
Lab 7 Osmosis
Time Required: 2 1/2 hours (plus 48 hours for preparation and observation)
Additional Materials: Cutting Board, 4 Eggs, Kitchen Knife, 2 Potatoes,
Stopwatch, Water
Lab 8 Enzymes
Time Required: 1 hour (plus 1 hour for preparation)
Additional Materials: Cutting Board, 2 Food Products, Hot Water Bath,
Kitchen Knife, Paper Towel, Saliva Sample, Stopwatch, Water Lab 9 Cellular Respiration
Time Required: 1 hour (plus 24 hours for preparation and time for overnight observations)
Additional Materials: Hot Water, Paper Towels, Stopwatch Lab 10 Cell Structure and Function
Time Required: 1 1/2 hours (plus 24 hours for observation)
Additional Materials: Bowl, 2 Household Items, Warm Water, Computer,
Internet Access Lab 11 Mitosis
Time Required: 3 hours
Additional Materials: None
8
Time and Materials
Lab 12 Meiosis
Time Required: 1 1/2 hours
Additional Materials: Computer, Internet Access Lab 13 DNA and RNA
Time Required: 2 hours
Additional Materials: Fruit, Scissors, Pen/Pencil Lab 14 Mendelian Genetics
Time Required: 1 1/2 hours
Additional Materials: None Lab 15 Population Genetics
Time Required: 1 1/2 hours
Additional Materials: Access to a Printer Lab 16 Taxonomy
Time Required: 1 hour
Additional Materials: None Lab 17 Bacteria and Archaea
Time Required: 1 hour (plus 24 hours preparation and 7 days for observation)
Additional Materials: Microwave or Hot Water Bath, Hot Pad, Refrigerator
9
Time and Materials
Lab 18 Protista
Time Required: 2 hours
Additional Materials: eScience Lab’s Student Portal Account, Internet
Access
Lab 19 Fungi
Time Required: 1 hour (plus 3 - 7 days for observation)
Additional Materials: White Bread, Water Lab 20 Energy and Photosynthesis
Time Required: 1 hour (plus 3 hours for observation)
Additional Materials: Cutting Board, Kitchen Knife, Light Source, Pencil,
Quarter, Scissors, Spinach Leaves (Fresh), Tape, Water Lab 21 Plant Circulation
Time Required: 1 1/2 hours (plus 24 hours for observation)
Additional Materials: 2 Celery Stalks, Cutting Board, Kitchen Knife, Water, Scissors, 3 Additional Environments Lab 22 Plant Reproduction
Time Required: 2 hours
Additional Materials: Fresh Flower (Lily is recommended) Lab 23 Invertebrates and Vertebrates
Time Required: 3 hours
Additional Materials: Paper Towel, Water
10
Time and Materials Lab 24 Animal Structure
Time Required: 1 hour (plus 7 additional days for observation)
Additional Materials: 1 Egg, Water
Lab 25 The Circulatory and Respiratory Systems
Time Required: 2 hours
Additional Materials: Water, Scissors
Lab 26 The Sensory and Nervous Systems
Time Required: 2 1/2 hours
Additional Materials: Pen/Pencil, Tape, White Construction or Printer
Paper, Water, Willing Participant
Lab 27 Ecology of Organisms
Time Required: 30 minutes (plus 7 days for observation)
Additional Materials: Paper Towel, Scissors, Sunny Location, Water
Lab 28 Ecological Interactions
Time Required: 30 minutes
Additional Materials: Large Bowl, Paper Towel, Large Pitcher, Water
11
Safety Information
Lab Safety eScience Labs, LLC designs every kit with safety as our top priority. Nonetheless, these are science kits and contain items which must be handled with care.
Safety in the laboratory always comes first!
Always follow the instructions in your laboratory manual and these general rules:
Lab Preparation
•
Please thoroughly read the lab exercise before starting!
•
If you have any doubt as to what you are supposed to be doing and how to do it safely, please STOP and then:
Double-check the manual instructions.
Check www.esciencelabs.com for updates and tips.
Contact us for technical support by phone at 1-888-ESL-Kits (1-888-375-5487) or by email at Help@esciencelabs.com.
•
Read and understand all labels on chemicals.
If you have any questions or concerns, refer to the Material Safely Data Sheets
(MSDS) available at www.esciencelabs.com. The MSDS lists the dangers, storage requirements, exposure treatment and disposal instructions for each chemical.
•
Consult your physician if you are pregnant, allergic to chemicals, or have other medical conditions that may require additional protective measures.
Proper Lab Attire
•
Remove all loose clothing (jackets, sweatshirts, etc.) and always wear closed-toe shoes. •
Long hair should be pulled back and secured and all jewelry (rings, watches, necklaces, earrings, bracelets, etc.) should be removed.
•
Safety glasses or goggles should be worn at all times. In addition, wearing soft contact lenses while conducting experiments is discouraged, as they can absorb potentially harmful chemicals.
•
When handling chemicals, always wear the protective goggles, gloves, and apron provided. 13
Safety Information
Performing the Experiment
•
Do not eat, drink, chew gum, apply cosmetics or smoke while conducting an experiment.
•
Work in a well ventilated area and monitor experiments at all times, unless instructed otherwise. •
When working with chemicals:
Never return unused chemicals to their original container to avoid contamination.
Never place chemicals in an unmarked container to avoid identification or proper disposal problems.
Always put lids back onto chemicals immediately after use to avoid contamination or potential hydration problems.
Never ingest chemicals. If this occurs, seek immediate help.
Call 911 or “Poison Control” 1-800-222-1222
•
Never pipette anything by mouth.
•
Never leave a heat source unattended.
If there is a fire, evacuate the room immediately and dial 911.
Lab Clean-up and Disposal
•
If a spill occurs, consult the MSDS to determine how to clean it up.
•
Never pick up broken glassware with your hands. Use a broom and a dustpan and discard in a safe area.
•
Do not use any part of the lab kit as a container for food.
•
Safely dispose of chemicals. If there are any special requirements for disposal, it will be noted in the lab manual.
•
When finished, wash hands and lab equipment thoroughly with soap and water.
Above all, use common sense! Read the manual carefully. Pay close attention to the safety concerns prior to starting an experiment.
14
Student Portal
Introduction:
Concept Animation
ESL Safety Video
ESL Scientific Processes Video
How Big Is It?
Introduction to the Microscope
Measuring Volume Using a
Graduated Cylinder
Unit Conversions
Lab Drill
Biological Processes:
Concept Animation
ESL Biological Processes Video
The Structure of an Atom
Acid/Base Reactions
Diffusion and Osmosis Tutorial
Docking Tutorial
Lab Drill
The Cell:
Concept Animation
Log on to the Student Portal using these easy steps:
Visit our website, www.eScienceLabs.com, and click on the button which says
“Register or Login” on the top right side of the homepage. From here, you will be taken to a login page. If you are registering your kit code for the first time, click the “Create an Account” hyperlink. Locate the kit-code, located on a label on the inside of the kit box lid. Enter this, along with other requested information into the online form to create your user account. Be sure to keep track of your username and password as this is how you will enter the Student Portal for future visits. This establishes your account with the eScience Labs’ Student Portal.
Have fun!
ESL Cell Video
Cell Structure Crossword Puzzle
Interactive Videos of Meiosis
Interactive Videos of Mitosis
Nature’s Review of RNA
DNA Transcription & Translation
How Mutations Work
Riken Center’s Developmental Biology Stem Cell Videos
A Typical Animal Cell
15
Student Portal
Construction of the Cell Membrane
The Cell Cycle
Cell Division
DNA Extraction Virtual Lab
Lab Drill
Kingdoms of Life:
Concept Animation
ESL Kingdoms Video
Biology of Bacteria
Biology of Flagellates
Biology of Algae
Protists Under Normal Conditions
Protists With Slowing Agent
Tree of Life Web Project
Virtual Pond Dip
Lab Drill
Plant Kingdom:
Concept Animation
ESL Plant Kingdom Video
Biology of Plants
Lab Drill
Animal Kingdom:
Concept Animation
ESL Animal Kingdom Video
Biology Crossword Puzzle
Biology of Echinoderms
Biology of Cnidarians
Biology of Chordates
Biology of Annelids
Anatomical Terminology: Body Regions
Anatomical Terminology: Relative Position
Regional Body Parts
16
Student Portal
The Skelton: Bones & Joints
Major Muscles of the Human Body
The Sense of Hearing
The Sense of Sight
The Sense of Smell
The Sense of Taste
The Anatomy of the Heart
Respiratory Basics
The Respiratory System
Lab Drill
Additional Resources:
Stop Watch
Conversion Tables
17
Sample Labware
19
Lab 1
Introduction to Science
Introduction to Science
Learning Objectives
•
Apply the scientific method, including making observations, developing hypotheses, identifying variables and controls, collecting and analyzing data, and drawing conclusions
•
Use calculations and measurements to connect percent error, significant figures, conversions, accuracy and precision to scientific reasoning
•
Review how to write and format a lab report
Introduction
What is science? You have likely taken several classes throughout your career as a student, and know that it is more than just chapters in a book. Science is a process. It uses evidence to understand the history of the natural world and how it works. Scientific knowledge is not static, but constantly evolving as we understand more about the natural world. Furthermore, the constant development of equipment and techniques allows us to gain an increasingly deeper insight into the natural world. Science begins with observations that can be measured in some way, and often concludes with observations from analyzed data.
Following the scientific method helps to minimize bias and increase validity when testing a theory. It helps scientists collect and organize information in a useful way so that patterns and data can be meaningfully analyzed. As a scientist, you should use the scientific method as you conduct the experiments throughout this manual. Figure 1: The scientific method process.
23
Introduction to Science
The process of the scientific method begins with an observation. For example, suppose you observe a plant growing towards a window. This observation could be the first step in designing an experiment! Remember that observations are used to begin the scientific method, but they may also be used to help analyze data.
Observations can be quantitative (measurable), or qualitative (immeasurable; observational). Quantitative observations allow us to record findings as data, and leave little room for subjective error. Qualitative observations cannot be measured. Instead, they rely on human sensory perceptions. The nature of these observations makes them more subjective and susceptible to human error. However, qualitative observations are still able to provide useful information.
For example, suppose you have a handful of pennies. You can make quantitative observations that there are
15 pennies, and each is 1.9 cm in diameter. Both the quantity, and the diameter, can be precisely measured.
You can also make qualitative observations that they are brown, shiny, and smooth. The color and texture are not numerically measured, and may vary based on the individual’s perception or background.
Quantitative observations are generally preferred in science because they involve "hard" data. Because of this, many scientific instruments, such as microscopes and scales, are used to alleviate the need for qualitative observations. Rather than observing that an object is large, scientists can identify specific mass, shapes, structures, etc.
Developing a Hypothesis
Once an observation has been made, the next step is to develop a hypothesis. A hypothesis is a testable statement describing what the scientist thinks will happen in the experiment. In other words, it is a proposed explanation for an event based on observation(s). For every hypothesis, a scientist also develops a null hypothesis. A null hypothesis is a testable statement that, if proven true, means the hypothesis was incorrect.
Both a hypothesis and a null hypothesis must be testable, but only one can be true.
Hypotheses are typically written in an if/then format. For example:
Hypothesis:
If plants are grown in soil with added nutrients, then they will grow faster than plants grown without added nutrients.
Null hypothesis:
If plants are grown in soil with added nutrients, then they will grow at the same rate as plants grown in soil without nutrients.
Figure 2: What affects plant growth?
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Introduction to Science
If plants grow quicker when nutrients are added, then the hypothesis is accepted and the null hypothesis is rejected.
Testing a Hypothesis
There are often many ways to test a hypothesis. However, three rules must always be followed during an experiment for results to be valid.
1. The experiment must be replicable.
2. Only one variable may be tested at a time
3. A control must always be included.
Replicability
Let’s begin by exploring the first rule. Experiments must be replicable to create valid theories. In other words, procedures must always be diligently recorded, and experiments must provide precise. Precise results are those which have very similar values (e.g., 85, 86, and 86.5) over multiple trials. By contrast, accurate results are those which demonstrate what you expected to happen (e.g., you expect amylase, an enzyme found in human saliva, to break down starch molecules during digestion).
Figure 3: Left: Accurate results all hit the bulls-eye on a target. Right: Precise results may not hit the bulls-eye, but they all hit the same region.
The following example demonstrates the significance of experimental replicability. Suppose you conduct an experiment and conclude that ice melts in 30 seconds when placed on a burner, but you do not record your procedure or define the variables. The conclusion that you draw will not be recognized in the scientific community because other scientists cannot repeat your experiment and find the same results. What if another scientist tries to repeat your ice experiment, but does not turn on the burner; or, uses a larger ice chunk than you used? The results will not be the same, because the experiment was not repeated using the same exact procedure. In order for results to be valid, repeated experiments must follow the original experiment exactly. Using this technique, multiple trials performed in this manner should yield comparable results.
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Introduction to Science
Variables
Variables are defined, measurable components of an experiment. Controlling variables in an experiment allows a scientist to quantify changes that occur. This allows for focused results to be measured; and, for refined conclusions to be drawn. There are two types of variables: independent and dependent variables.
Independent variables are variables that scientists select to change within the experiment. For example, the time of day, amount of substrate, etc. can all be independent variables. Independent variables are also used by scientists to develop hypotheses. The “if” part of the hypothesis describes the independent variable and how the scientist will manipulate it. For example, the independent variable in the hypothesis, “If plants are grown in soil with added nutrients, then they will grow faster than plants grown without added nutrients,” is soil with added nutrients. Experiments can only have one independent variable. In this way, scientists can determine if altering the independent variable is the reason for obtaining a different result. Scientists would not be able to conclusively determine which change affected the data if more than one independent variable is changed in an experiment. However, a hypothesis is usually developed to focus on one variable only. The dependent variable is reflected in the “then” part of the hypothesis. For example, if there is a change in the independent variable, then the dependent variable will also change. Independent variables are always placed on the x-axis of a chart or graph.
Dependent variables are variables which are observed in relationship to the independent variable. Any changes observed in the dependent variable are caused by the changes in the independent variable. In other words, they depend on the independent variable. Common examples of this are: reaction rate, color change, etc. There can be more than one dependent variable in an experiment. Dependent variables are placed on the y-axis of a chart or graph.
Controls
A control is a sample of data collected in an experiment that is not exposed to the independent variable. The control sample reflects the factors that could influence the results of the experiment, but do not reflect the planned changes that might result from manipulating the independent variable. Controls must be identified to eliminate compounding changes that could influence results. Often, the hardest part of designing an experiment is determining how to isolate the independent variable and control all other possible variables. Scientists must be careful not to eliminate or create a factor that could skew the results. For this reason, taking notes to account for unidentified variables is important. This might include factors such as temperature, humidity, time of day, or other environmental conditions that may impact results.
There are two types of controls, positive and negative. Negative controls are data samples in which you expect no change to occur. They help scientists determine that the experimental results are due to the independent variable, rather than an unidentified or unaccounted variable. For example, suppose you need to culture (grow) bacteria and want to include a negative control. You could create this by streaking a sterile loop across an agar plate. Sterile loops should not create any microbial growth; therefore, you expect no change to occur on the agar plate. If no growth occurs, you can assume the equipment used was sterile. However, if
26
Introduction to Science microbial growth does occur, you must assume that the equipment was contaminated prior to the experiment and must redo the experiment with new materials.
Alternatively, positive controls are data samples in which you do expect a change. Let’s return to the growth example, but now you need to create a positive control. To do this, you now use a sterile loop to streak a plate with a bacterial sample that you know grows well on agar (such as E. coli). If bacteria grows, you can assume that the bacteria sample and agar are both suitable for the experiment. However, if bacteria does not grow, you must assume that the agar or bacteria has been compromised; the agar is inhibiting growth, or the bacteria in the sample are not viable.
Collecting and Presenting Data
The scientific method also requires data collection. This may reflect what occurred before, during, or after an experiment. Collected data help reveal experimental results. Data should include all relevant observations, both quantitative and qualitative.
After results are collected, they can be analyzed. Data analysis often involves a variety of calculations, conversions, graphs, tables, etc. A common task a scientist faces is unit conversion. Units of time are often displayed in an increment that must be converted. For example, suppose half of your data is measured in seconds, but the other half is measured in minutes. It will be difficult to understand the relationship between the data if the units are not equivalent (sample calculation below).
Significant Digits
When calculating a unit conversion, significant digits must be accounted for. Significant digits are the digits in a number or answer that describe how precise the value actually is. Consider the rules in Table 1.
Addition and subtraction problems should result in an answer that has the same number of significant decimal places as the least precise number in the calculation. Multiplication and division problems should keep the same total number of significant digits as the least precise number in the calculation. For example:
Addition Problem: 12.689 + 5.2 = 17.889
Multiplication Problem: 28.8 x 54.76 = 1577.088
round to 18
round to 1580 (3 significant digits)
Scientific notation is another common method used to report a number. Scientific data is often very large
(e.g., the speed of light) or very small (e.g., the diameter of a cell). Scientific notation provides an abbreviated expression of a number, so that scientists don’t get caught up counting a long series of zeroes.
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Introduction to Science
Table 1: Significant Digits Rules
Rule
Examples
•
45 has two significant digits
•
3.99 has three significant digits
Any time a zero appears between significant numbers, the zero is significant.
•
4005 has four significant digits
•
0.3400000009 has ten significant digits
Zeros that are ending numbers after a decimal point or zeros that are after significant numbers before a decimal point are significant.
•
45.00 has four significant digits
•
15000.00 has seven significant digits
Zeros that are used as placeholders before other digits are NOT significant digits.
•
.0000000897 has three significant digits
A zero at the end of a number with no decimal can be a significant digit.
*To avoid uncertainty, numbers can be written using scientific notation.
•
6200 can have 2, 3, or 4 significant digits
(e.g., 6.2 x 104 has 2, 6.20 x 104 has 3, and
6.200 x 104 has 4)
Any non-zero number (1 - 9) is always significant.
There are three parts to scientific notation: the base, the coefficient and the exponent. Base 10 is almost always used and makes the notation easy to translate. The coefficient is always a number between 1 and 10, and uses the significant digits of the original number. The exponent tells us whether the number is greater or less than 1, and can be used to “count” the number of digits the decimal must be moved to translate the number to regular notation. A negative exponent tells you to move the decimal to the left, while a positive one tells you to move it to the right.
Figure 4: The exponent equals the number of decimal places moved until the co- For example, the number 5,600,000 can be written in scientific noefficient is a number between 1 and 10.
6
tation as 5.6 x 10 . The coefficient is 5.6, the base is 10, and the exponent is 6. If you multiply 5.6 by 10 six times, you will arrive at 5,600,000. Note the exponent, 6, is positive because the number is larger than one. Alternative, the number 0.00045 must be written using a negative exponent. To write this number in scientific notation, determine the coefficient. Remember that the coefficient must be between 1 and 10. The significant digits are 4 and 5. Therefore, 4.5 is the coefficient. To determine the exponent, count how many places you must move the decimal over to create the original number. Moving to the left, we have 0.45, 0.045, 0.0045, and finally 0.00045. Since we move the decimal 4 places to the left, the exponent is -4. Written in scientific notation, we have 4.5 x 10-4
Although these calculations may feel laborious, a well-calculated presentation can transform data into a format that scientists can more easily understand and learn from. Some of the most common methods of data presentation are tables and graphs.
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Introduction to Science
Table: A well-organized summary of data collected. Tables should display any information relevant to the hypothesis. Always include a clearly stated title, labeled columns and rows, and measurement units.
Table 2: Plant Growth With and Without Added Nutrients
Variable
Height Wk. 1 (mm)
Height Wk. 2 (mm)
Height Wk. 3 (mm)
Height Wk. 4 (mm)
Control
3.4
3.6
3.7
4.0
3.5
3.7
4.1
4.6
(without nutrients)
Independent
(with nutrients)
Graph: A visual representation of the relationship between the independent and dependent variable. Graphs are useful in identifying trends and illustrating findings. Rules to remember:
•
The independent variable is always graphed on the x-axis (horizontal), with the dependent variable on the y axis (vertical).
•
Use appropriate numerical spacing when plotting the graph, with the lower numbers starting on both the lower and left hand corners.
•
Always use uniform or logarithmic intervals. For example, if you begin by numbering, 0, 10, 20, do not jump to 25 then to 32.
•
Title the graph and both the x and y axes such that they correspond to the data table from which they come. For example, if you titled your table “Heart rate of those who eat vegetables and those who do not eat vegetables”, the graph title should reference this information as well.
•
Determine the most appropriate type of graph. Typically, line and bar graphs are the most common. Line Graph: Shows the relationship between variables using plotted points that are connected with a line. There must be a direct relationship and dependence between each point connected.
More than one set of data can be presented on a line graph. Figure 5 uses the data from
Table 2.
Plant growth, with and without nutrients, over time.
Figure 5: Sample line graph.
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Introduction to Science
Speed (kph)
Bar Graph: Used to compare results that are independent from each other, as opposed to a continuous series. Since the results from our previous example are continuous, they are not appropriate for a bar graph.
Figure 6: Sample bar graph. Top speed for Cars A, B, C, and D.
After compiling the data, scientists analyze the data to determine if the experiment supports or refutes the hypothesis. If the hypothesis is supported, you may want to consider additional variables that should be examined. If your data does not provide clear results, you may want to consider running additional trials or revising the procedure to create a more precise outcome.
Percent Error
One way to analyze data is to calculate percent error. Many experiments perform trials which calculate known values. When this happens, you can compare experimental results to known values and calculate percent error. Low percent error (<5%) indicates that results are probably accurate, and high percent error
(>20%) indicates that results may be inaccurate. The formula for percent error is:
Percent Error = |(Experimental - Actual)| x 100%
Actual
Note that the brackets flanking the numerator indicate “absolute value”. This means that the number in the equation is always positive.
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Introduction to Science
Suppose your experiment involves gravity. Your experimental results indicate that the speed of gravity is
10.1 m/s2, but the known value for gravity is 9.8 m/s2. We can calculate the percent error through the following steps:
Percent Error = |(10.1 m/s2 - 9.8 m/s2)|
x 100%
(9.8 m/s2)
Percent Error = |0.3|
x 100%
(Note the units cancel each other out)
(9.8)
Percent Error = 0.0306 x 100% = 3.1% (Remember the significant digits)
Writing a Lab Report
The scientific method gives us a great foundation to conduct scientific reasoning. The more data and observations we are able to make, the more we are able to accurately reason through the natural phenomena which occur in our daily lives. Scientific reasoning does not always include a structured lab report, but it always helps society to think through difficult concepts and determine solutions. For example, scientific reasoning can be used to create a response to the changing global climate, develop medical solutions to health concerns, or even learn about subatomic particles and tendencies.
Although the scientific method and scientific reasoning can guide society through critical or abstract thinking, the scientific industry typically promotes lab reports as a universal method of data analysis and presentation. In general terms, a lab report is a scientific paper describing the premise of an experiment, the procedures taken, and the results of the study. They provide a written record of what took place to help others learn and expedite future experimental processes.
Though most lab reports go unpublished, it is important to write a report that accurately characterizes the experiment performed.
Figure 7: Lab reports are an important part of science, providing a way to report conclusions and ideas.
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Introduction to Science
Table 3 summarizes the components of a typical lab report.
Table 3: Lab Report Components
Lab Report
Purpose
Section
Title
Abstract
A short statement summarizing the topic
A brief summary of the methods, results and conclusions. It should not exceed
200 words and should be the last part written.
An overview of why the experiment was conducted. It should include:
Introduction
•
Background - Provide an overview of what is already known and what questions remain unresolved. Be sure the reader is given enough information to know why and how the experiment was performed.
•
Objective - Explain the purpose of the experiment (i.e. "I want to determine if taking baby aspirin every day prevents second heart attacks.")
•
Hypothesis - This is your "guess" as to what will happen when you do the experiment. Materials and
Methods
A detailed description of what was used to conduct the experiment, what was actually done (step by step) and how it was done. The description should be exact enough that someone reading the report can replicate the experiment.
Results
Data and observations obtained during the experiment. This section should be clear and concise. Tables and graphs are often appropriate in this section. Interpretations should not be included here.
Data interpretations and experimental conclusions.
Discussion
32
•
Discuss the meaning of your findings. Look for common themes, relationships and points that perhaps generate more questions.
•
When appropriate, discuss outside factors (i.e. temperature, time of day, etc.) that may have played a role in the experiment.
•
Identify what could be done to control for these factors in future experiments.
Conclusion
A short, concise summary that states what has been learned.
References
Any articles, books, magazines, interviews, newspapers, etc. that were used to support your background, experimental protocols, discussions and conclusions.
Introduction to Science
Exercise 1: Data Interpretation
Dissolved oxygen is oxygen that is trapped in a fluid, such as water. Since many living organism requires oxygen to survive, it is a necessary component of water systems such as streams, lakes and rivers in order to support aquatic life. The dissolved oxygen is measured in units of ppm (parts per million). Examine the data in
Table 4 showing the amount of dissolved oxygen present and the number of fish observed in the body of water the sample was taken from; finally, answer the questions below.
Table 4: Water Quality vs. Fish Population
Dissolved Oxygen (ppm)
0
2
4
6
8
10
12
14
16
18
Number of Fish Observed
0
1
3
10 12
13
15
10
12
13
1. What patterns do you observe based on the information in Table 4?
2. Develop a hypothesis relating to the amount of dissolved oxygen measured in the water sample and the number of fish observed in the body of water.
3. What would your experimental approach be to test this hypothesis?
4. What would be the independent and dependent variables?
5. What would be your control?
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Introduction to Science
6. What type of graph would be appropriate for this data set? Why?
7. Graph the data from Table 4: Water Quality vs. Fish Population (found at the beginning of this exercise).
8. Interpret the data from the graph made in Question 7.
Exercise 2: Testable Observations
Determine which of the following observations are testable. For those that are testable:
Determine if the observation is qualitative or quantitative
Write a hypothesis and null hypothesis
What would be your experimental approach?
What are the dependent and independent variables?
What are your controls - both positive and negative?
How will you collect your data?
How will you present your data (charts, graphs, types)?
How will you analyze your data?
Observations
1. A plant grows three inches faster per day when placed on a window sill than it does when placed on a on a coffee table in the middle of the living room.
2. The teller at the bank with brown hair and brown eyes is taller than the other tellers.
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Introduction to Science
3. When Sally eats healthy foods and exercises regularly, her blood pressure is 10 points lower than when she does not exercise and eats fatty foods.
4. The Italian restaurant across the street closes at 9 pm but the one two blocks away closes at 10 pm.
5. For the past two days, the clouds have come out at 3 pm and it has started raining at 3:15 pm.
6. George did not sleep at all the night following the start of daylight savings.
Exercise 3: Conversion
For each of the following, convert each value into the designated units. 1. 46,756,790 mg = _______ kg
2. 5.6 hours = ________ seconds
3. 13.5 cm = ________ inches
4. 47 °C = _______ °F
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Introduction to Science
Exercise 4: Accuracy and Precision
For the following, determine whether the information is accurate, precise, both or neither.
1. During gym class, four students decided to see if they could beat the norm of 45 sit-ups in a minute. The first student did 64 sit-ups, the second did 69, the third did 65, and the fourth did 67.
2. The average score for the 5th grade math test is 89.5. The top 5th graders took the test and scored 89,
93, 91 and 87.
3. Yesterday the temperature was 89 °F, tomorrow it’s supposed to be 88 °F and the next day it’s supposed to be 90 °F, even though the average for September is only 75 °F degrees!
4. Four friends decided to go out and play horseshoes. They took a picture of their results shown to the right:
5. A local grocery store was holding a contest to see who could most closely guess the number of pennies that they had inside a large jar. The first six people guessed the numbers 735, 209, 390, 300, 1005 and
689. The grocery clerk said the jar actually contains 568 pennies.
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Introduction to Science
Exercise 5: Significant Digits and Scientific Notation
Part 1: Determine the number of significant digits in each number and write out the specific significant digits.
1. 405000
2. 0.0098
3. 39.999999
4. 13.00
5. 80,000,089
6. 55,430.00
7. 0.000033
8. 620.03080
Part 2: Write the numbers below in scientific notation, incorporating what you know about significant digits.
1. 70,000,000,000
2. 0.000000048
3. 67,890,000
4. 70,500
5. 450,900,800
6. 0.009045
7. 0.023
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Introduction to Science
Exercise 6: Percentage Error
In the questions below, determine the percentage error. Show your work on all problems.
1. A dad holds five coins in his hand. He tells his son that if he can guess the amount of money he is holding within 5% error he can have the money. The son guesses that he is holding 81 cents. The dad opens his hand and displays 90 cents. Did the son guess close enough to receive the money from his father? 2. A science teacher tells her class that their final project requires the students to measure a specific variable and determine the velocity of a car with no more than 2.5% error. Jennifer and Johnny work hard and decide the velocity of the car is 34.87 m/s. The teacher informs them that the actual velocity is 34.15 m/s. Will Jennifer and Johnny pass their final project?
3. A locomotive train is on its way from Chicago, IL to Madison, WI. The trip is said to last 3.15 hours. When the train arrives in Madison the conductor notices it actually took them 3.26 hours. The train company prides itself on always having its trains to the station within a 3% error of the expected time. Will the train company live up to its reputation on this trip?
4. A coach tells his little league players that hitting a 0.275 batting average, within 7% percentage error, means that they had a really great season. Seven year old Tommy ended the season hitting a 0.258 batting average. According to his coach, did he have a great season?
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Introduction to Science
Exercise 7: Experimental Variables
Determine the variables tested in the each of the following experiments. If applicable, determine and identify any positive or negative controls.
1. A study is being done to test the affects of habitat space on the size of fish populations. Different sized aquariums are set up with six goldfish in each one. Over a period of six months, the fish are fed the same type and amount of food. The aquariums are equally maintained and cleaned throughout the experiment.
The temperature of the water is kept constant. At the end of the experiment the number of surviving fish are surveyed.
A. Independent Variable:
B. Dependent Variable:
C. Controlled Variables/Constants:
D. Experimental Controls/Control Groups:
2. To determine if the type of agar affects bacterial growth, a scientist cultures E. coli on four different types of agar. Five petri dishes are set up to collect results:
•
One with nutrient agar and E. coli
•
One with mannitol-salt agar and E. coli
•
One with MacConkey agar and E. coli
•
One with LB agar and E. coli
•
One with nutrient agar but NO E. coli
All of the petri dishes received the same volume of agar, and were the same shape and size. During the experiment, the temperature at which the petri dishes were stored, and at the air quality remained the same.
After one week the amount of bacterial growth was measured.
A. Independent Variable:
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Introduction to Science
B. Dependent Variable:
C. Controlled Variables/Constants:
D. Experimental Controls/Control Groups:
40
Lab 2
General Lab Safety
General Lab Safety
Learning Objectives
•
Explain how to safely work in a biological and chemical laboratory
•
Explain how and when to use the safety equipment in a biological and chemical laboratory
•
Identify common laboratory equipment
•
Properly neutralize and dispose of acidic and basic solutions
Introduction
If you’ve ever seen the Frankenstein movies, then you are probably familiar with Hollywood’s interpretation of a laboratory as a dark, dungeonesque space with a mad scientist in action. However, although lab experiences may often appear haphazard in movies, real-world scientists develop and incorporate organized protocols to keep lab environments safe. In addition to focusing on safe lab practices, the experiments in this lab manual will help you to learn about “green”, environmentally-friendly chemicals and adhere to safety precautions.
Before beginning these labs, you must first realize there are specific techniques and precautions to learn to decrease the risks involved. You will need to learn the location of certain items, rules, and the names of common lab equipment. After you get acquainted with your surroundings, you will be prepared and comfortable completing the labs in this manual.
This manual is written for a “Green Chemistry” approach. This means the activities in this manual are generally safer and produce less hazardous waste than similar experiments found in other lab manuals. The chemicals used are environmentally friendly and can be discarded down a household drain with running water. Each of these experiments has been performed safely by other students. However, safer and less hazardous waste does not mean accidents, injuries, or damage will not happen. Scrapes and burns are always a possibility. These experiments will likely be performed in a household environment, so be sure a working fire extinguisher is accessible. You are advised to always use the safety glasses and gloves provided in your kit. Note that the gloves provided in your lab kit are nitrile-based, rather than latex. Please contact eScience
Labs at 1-888-ESL-KITS or info@eScienceLabs.com if you require and alternate.
Safety Equipment
• Safety Glasses: Safety glasses are used to protect your eyes, and should be worn at all times when in the chemistry laboratory, even if you are not currently working with chemicals.
•
Gloves: At times you may need to wear gloves to protect your hands from harmful chemicals or hot objects. The type of glove needed will depend on the application. For example, oven mitts are worn to remove hot objects from an oven while vinyl or latex gloves are used when working with
43
General Lab Safety acids and bases. It is very important not to touch your work area with gloves that have been contaminated with harmful chemicals.
•
Safety Shower: A safety shower is used when a hazardous chemical is spilled on a person where they are unable to rinse it off thoroughly in the sink. It can also be used if a person’s clothes catch on fire. Most safety showers are operated by pulling a chain, although a standard home shower will also work. If a sink is insufficient to thoroughly rinse yourself, get to the nearest shower in your home as quickly as possible. This is not a time for modesty. Remove the contaminated clothing while rinsing your skin with a copious amount of water.
Figure 1: Using the information in this section, you will be able to complete the labs experiments in this manual safely and successfully. Can you identify what’s wrong with this picture?
•
Eye Wash: An eye wash is used if a harmful chemical is splashed into your eyes or face. It is usually operated by pushing forward on a handle. In the home, find the nearest sink and flush water in your eyes for at least 20 minutes. If your sink has a sprayer, use it to rinse your eyes or face while making sure the water drains into the sink.
•
Fire Extinguisher: A fire extinguisher is used to put out small to medium fires.
•
Laboratory Fume Hood: A laboratory fume hood removes harmful gases and fumes sometimes present when doing an experiment. You should always work in a fume hood whenever you are working with corrosive, noxious, or flammable materials. Chemicals used in this kit will not require the use of a fume hood.
Besides knowing where the safety equipment is located and how and when to use it, there are general safety rules you need to follow in the laboratory. Some of the common safety rules are listed below.
Laboratory Safety Rules
44
1.
Always wear safety glasses. Never wear contact lenses.
2.
Never attempt unauthorized experiments.
3.
Always have someone available to help in the event of an accident.
4.
Never have food, drink, chewing gum, or tobacco in the laboratory.
5.
Always keep your work area free of clutter.
6.
Always wear a protective apron and sensible clothing. This means no loose clothing, bare midriffs, or open-toe shoes.
7.
Know the location of and how to use safety equipment in your home. This includes showers, fire
General Lab Safety extinguishers, and sinks.
8.
Always read the experiment entirely before beginning the procedure.
9.
Always wash hands before leaving the lab.
10. Tie back long hair.
11. Never run or play practical jokes in the experiment area.
12. Place broken glass in a protective container, never loosely in a trash can.
Acid-Base Behavior
In addition to following the general safety rules, chemicals need to be handled properly. In particular, two very important classes of compounds called acids and bases require special attention. These compounds are commonly used reagents in the laboratory; therefore, understanding their proper disposal is beneficial.
Physical differences between acids and bases can be detected by the some of the five senses, including taste and touch. Acids have a sour or tart taste and can produce a stinging sensation to broken skin. For example, lemon juice tastes sour. Alternatively, bases have a bitter taste and a slippery texture. Soap and many cleaning products are bases.
Acids and bases are measured on a scale called pH. pH, or potential hydrogen, is calculated using a mathematical equation that accounts for some of Table 1: pH Range of Common
Foods
the chemical differences in acidic and basic compounds. This scale helps
Food
pH Range us quickly determine if a solution is very acidic, a little acidic, neutral
(neither acidic nor basic), a little basic, or very basic. pH values range
Lime
1.8 - 2.0 from less than 1 to 14. A pH of 1 is highly acidic, a pH of 14 is highly basic
(or alkaline), and a pH of 7 is neutral. Table 1 lists the pH of several foods. pH indicators, which change color under a certain pH level, can be used to determine whether a solution is acidic or basic. For example, litmus paper is made by coating a piece of paper with litmus, which changes color at around a pH of 7. Either red or blue litmus paper can be purchased depending on the experimental needs. Blue litmus paper remains blue when dipped in a base, but turns red when dipped in an acid, while red litmus paper stays red when dipped in an acid, but turns blue when in contact with a base.
Acids and bases can react with each other. In this case, the two opposites cancel each other out resulting in a product that is neither an acid nor a
Soft Drinks
2.0 - 4.0
Apple
3.3 - 3.9
Tomato
4.3 - 4.9
Cheese
4.8 - 6.4
Potato
5.6 - 6.0
Drinking
Water
6.5 - 8.0
Tea
7.2
Eggs
7.6 - 8.0
45
General Lab Safety base. This type of reaction is called a neutralization reaction. Neutralization of an acid or base is a technique frequently used for the proper disposal of the compound. The neutralized product can usually be disposed of by flushing it down a sink.
Keep in mind that in a typical laboratory strong acids and bases, such as hydrochloric acid (HCl) and sodium hydroxide (NaOH), are commonly utilized. As mentioned previously, the experiments in this lab manual are designed with a green approach. That means that the acids and bases, in addition to the other chemicals classes, used in this manual are safe for direct disposal.
Some general guidelines for handling chemicals properly are listed below.
Handling Chemicals
46
1.
Always add acids to water, never water to acids.
2.
Never return unused chemicals to the bottles from where they were first obtained.
3.
Dispose of used chemicals in the proper waste containers and/or as instructed.
4.
Always clean the work area, and put away extra equipment when laboratory work is completed.
5.
Never leave anything unattended while it is being heated or is reacting rapidly.
6.
Never carry out a reaction or heat a substance in a closed system.
7.
Always be careful when working with previously heated objects.
8.
Always replace stoppers or lids on bottles containing chemicals.
9.
Weigh chemicals in weigh boats or on paper provided for that purpose. Never weigh chemicals by placing them directly on the scale.
10.
Label all chemicals clearly and completely.
11.
Read labels carefully before using chemicals.
12.
Always lubricate glass tubing or thermometers before inserting them into rubber stoppers.
13.
Material Safety Data Sheets (MSDS) for all chemicals provided can be found on our website at www.eScienceLabs.com/educators/msds. These sheets contain all needed information regarding the danger, safety and disposal of every chemical. Download, print, and review the MSDS for each chemical prior to using that chemical.
General Lab Safety
Even though you follow all of the safety rules, accidents can still happen. This is why it is so important to
know what to do for each type of accident.
How to Respond to Accidents
1. Chemical Spills (on the Bench or Floor): Be sure to clean up the spill immediately. If the spill involves volatile or flammable materials, such as alcohol, make sure ALL flames in the lab area are extinguished and spark-producing equipment is shutdown. In the case of an acid spill, pour baking soda on the acid before cleaning up. In the case of a base spill, pour vinegar on the base before cleaning it up. All other chemicals used in this manual can be cleaned up as you normally would. If you have any questions, check the MSDS.
2. Hazardous Chemical Spills (on a Person): If the spill covers a large area, the typical course of action is to remove all contaminated clothing while the person is under the safety shower. If it is a small area, flush the area immediately with a large amount of water and then wash it with soap. Check the MSDS for the spilled chemical and follow all instructions. Medical assistance may be necessary.
3. Chemicals Spills (in the Eyes): If a harmful chemical is splashed on your face and/or in your eyes, immediate attention is critical. Call for help and get to the nearest sink. If the chemical splashes on your face, and you have glasses on, KEEP the glasses on. Remove the chemical from your face before you remove the glasses. If a chemical gets in your eyes, hold your eyes open in the eyewash for at least 20 minutes. Even though you should not be wearing contact lenses in the lab, if you are, rinse your eyes thoroughly, remove your contacts, and continue to rinse your eyes. A doctor should examine your eyes as soon as possible.
4. Chemical Ingestion: Check the MSDS immediately. Call 911 or “Poison Help” at 1-800-222-1222.
5. Burns: Flush the area with cool running water for 20 minutes. Medical attention may be necessary.
6. Cuts and Wounds: If a chemical gets into the cut or wound, rinse it off immediately with a large amount of water. Avoid contamination, and check the MSDS.
7. Fire: Fires in a laboratory are often contained in pieces of glassware, such as a beaker. You should not move a beaker that has a chemical burning in it. Instead, extinguish the fire simply by covering the mouth of the beaker with a thin curved piece of glass called a watch glass and turning off the source of the flame. A plate or pie pan can also work in place of a watch glass. If the fire is not easily covered, you can use a fire extinguisher. If the fire is too large to extinguish quickly, clear the home and call the fire department immediately.
•
Clothing fires can be extinguished in a safety shower if it is close by. If it is not very close, you will need to STOP, DROP, and ROLL to quickly smother the fire.
47
General Lab Safety
Additional Resources
If you have any doubt as to what you are supposed to be doing and how to do it safely:
•
STOP!
•
Double-check the manual
•
Check the website www.esciencelabs.com
•
Email: Help@esciencelabs.com
•
Call for help 1-888-ESL-KITS (1-888-375-5487)
•
Contact your instructor
If you have ANY questions or concerns regarding a chemical, read the MSDS for that chemical. The MSDS lists the dangers, storage requirements, exposure treatment and disposal instructions for every chemical.
The MSDS for any chemical supplied by eScience Labs, LLC, can be found at www.eScienceLabs.com/ educators/msds. Pre-Lab Questions
1. What should you always wear to protect your eyes when you are in the chemistry laboratory?
2. Should you add acid to water or water to acid?
3. Where should you dispose of broken glass?
What should you do if you spill a chemical on your hand?
48
General Lab Safety
Exercise 1: What Is It?
A chemical laboratory contains special equipment to use while you are performing an experiment. Locate each of the items pictured on the following pages in your lab kit, and place a check mark in the appropriate place when you find it. After you have completed this, sketch a picture and name any additional items that are located in your lab kit, classroom, or home that are likely to be useful for you in completing these labs.
Beaker
50 mL
250 mL
S r S ck
Graduated Cylinder
10 mL
100 mL
Petri Dish
Test Tube
Pipette
49
General Lab Safety
Include your Drawings Here:
50
General Lab Safety
Experiment 1: Neutralization of Acids and Bases
In this experiment, you will learn how to properly neutralize and dispose of acidic and basic solutions.
Materials
5 mL 4.5% Acetic Acid (vinegar), C2H4O2
4 Weigh Boats
(1) 10 mL Graduated Cylinder
*Water
8 Litmus Test Strips (Neutral)
Permanent Marker
*You Must Provide
2 Pipettes
1 g Sodium Bicarbonate (baking soda), NaHCO3
Procedure
1. Use the permanent marker to label three of the weigh boats as A - C.
2. Measure and pour approximately 5 mL of water into weigh boat “A”.
3. Add 0.5 g sodium bicarbonate to weigh boat “B”.
4. Measure and pour approximately 5 mL of water into weigh boat “B”. Gently pipette the solution up and down until the sodium bicarbonate is fully dissolved in the water.
5. Measure and pour 5 mL acetic acid solution to weigh boat “C”.
6. Use the litmus test strips to determine if the substances in weigh boats A - C are acidic or basic. This is accomplished by briefly dipping an unused strip of the litmus paper in each of the weigh boats. Record your color results in Table 2.
7. Pipette 1 mL of the sodium bicarbonate solution from weigh boat “B” into weigh boat “C”. Gently swirl weigh boat “C” to mix.
8. Develop and record a hypothesis regarding the pH of weigh boat “C”. Record this in the Post-Lab Questions section.
9. Test the pH of weigh boat “C” using new litmus paper. Record your result in Table 3.
10. Repeat Step 9 four more times until all the sodium bicarbonate has been added to weigh boat “C”.
51
General Lab Safety
Table 2: Initial Litmus Test Results
Weigh Boat
Chemical Contents
Litmus Results
Additional Observations
A
B
C
Table 3: Neutralization of an Acid
Amount of Base
Litmus Result
1 mL
2 mL
3 mL
4 mL
5 mL
Post-Lab Questions
1. State your hypothesis (developed in Step 8) here. Be sure to include what you think the pH will be, and why. 2. What is a neutralization reaction?
3. When might neutralization reactions be used in a laboratory setting?
4. At what point was the acetic acid in weigh boat “C” neutralized?
5. What do you think would have been the results if a stronger solution of sodium bicarbonate was used?
Would it take more or less to neutralize the acid? What about a weaker concentration of sodium bicarbonate?
52
Lab 3
Chemical Bonding Fundamentals
Molecule Pictured: Arginine
Chemical Bonding Fundamentals
Learning Objectives
•
Relate matter to atoms, protons, electrons, and neutrons
•
Explain how electron configuration affects the interaction of atoms
•
Identify patterns in the periodic table of elements
•
Compare and contrast covalent and ionic bonding
•
Explain how covalent bonding can result in polar and non-polar molecules
Introduction
A myriad of reactions are constantly occurring within each living organism. These reactions allow an organism to reproduce, grow, move, eat, and perform a great many more functions. In order to properly study these biological reactions, a general understanding of chemistry is necessary.
The term matter is used to describe anything that occupies space and has mass. Take a look around, everything you can see and touch is matter (including you). The atomic theory states that all matter is made of atoms. Although physicists and other scientists have developed ways to split atoms to produce nuclear energy, atoms are considered the smallest indivisible unit of all matter. The word atom, in fact, comes from the
Greek word atomos meaning ‘cannot be cut’ or ‘indivisible’.
Atomic Structure
Atoms contain three types of subatomic particles: electrons (negatively charged particles), protons
(positively charged particles) and neutrons (uncharged, neutral particles). Within an atom, the subatomic particles are arranged so that the proton(s) and neutron(s) form a nucleus. The electrons orbit around the nucleus in a very specific manner. This atomic structure is analogous to the manner in which the planets orbit around the sun in our solar system. In this example, the nucleus represents the sun and the electrons represent the orbiting planets.
Instead of gravity holding the atom together, the positive charge of the protons attracts the negative electrons in an electrostatic manner. The configuration of electrons associated with an atom determines the chemical properties of that atom. Having a basic understanding of electron con- Figure 1: Diatomic oxygen molecules share four figuration aids in the overall comprehension of how at- valence electrons through covalent bonding. oms interact. As mentioned above, electrons are found in Note the electron orbitals and the proton-nucleus orbits that surround the nucleus. There are specific orbits bundle located near the center of each atom.
55
Chemical Bonding Fundamentals
(or “shells”) that contain electrons. These shells get filled from the innermost (closest to the nucleus) to the outermost shell. An atom is most stable (or, unreactive) when the outer shell is filled. Some atoms, such as helium, have a filled outer shell. Because of this, those atoms are not reactive. Most atoms do not have filled outer shells. In order to fill their outer shell, atoms can either gain or lose electrons to other atoms, or they can share electrons with other atoms.
Elements are pure substances that are made of only one type of atom. For example, gold is only made up of gold atoms. There are over 100 known elements, each with Figure 2: Cross-section of an atom. different chemical and physical properties. Interestingly, the number of naturally occurring elements is unclear as elements continue to be developed and/or detected in laboratories. The periodic table (Figure 4) has been used to categorize elements.
Figure 3: Elemental information for carbon.
The periodic table contains almost endless useful information for performing biology and chemistry. The periodic table is organized similar to a typical data table. It contains individual cells (compartmentalized squares) that are arranged logically in rows and columns according to specific trends. Each cell contains information about a particular element. In the middle of the cell is a one or two letter abbreviation for a particular element called a chemical symbol. For example, Au is the symbol for gold and Na is the symbol for sodium. Above an element’s symbol is the atomic number, which is the number of protons that exist in an atom of that element. Each element has its own unique number of protons. For example, gold atoms have 79 protons and sodium atoms have 11. The number of electrons is normally equal to the number of protons.
Therefore, the number of electrons in a gold atom is 79 and the number of electrons in sodium atoms is 11.
As you go across a row (called a period) moving left to right in the periodic table, you will find that the atomic number of the elements increases by one. This means the number of protons in the elements increases by one. The addition of that single proton changes the properties of the atom. On the other hand, when moving down a column (called a group) of the periodic table you will find that the number of electrons in the outer shell remains the same. This is why the atoms within a group have similar chemical behavior.
56
Chemical Bonding Fundamentals
Color Key:
Figure 4: The periodic table of elements categorizes all of the known elements. Groups are listed vertically as 1 7. Periods are listed horizontally as 1 - 18.
Chemical Bonding
In nature, most elements are not found alone; atoms of most elements combine to make molecules. A molecule is a mixture of two or more atoms in definite proportions. If the molecule contains atoms of different elements it is called a compound. These atoms are held together by chemical bonds, bringing them to a stable state. Keep in mind that a stable state is accomplished by the filling of the atoms’ outermost shell. Electrons involved with the chemical bonds between atoms are called valence electrons.
The two most common bonds are covalent bonds and ionic bonds. Covalent bonds form when two atoms share valence electrons. Ionic bonds form when an atom or molecule carries an electrical charge, which attracts an atom or molecule of the opposite charge. This electrical charge is created when an atom gains or loses electrons in an attempt to fill its outer shell. The resulting ions have opposite charges and attract one another via electrostatic attractions. Ionic and covalent bonds have very different physical properties. For example, ionic compounds have higher melting and boiling points. Ionic compounds also tend to be electrolytes. This means that they contain free ions which make them electrically conductive.
Ionic compounds are formed when anions (negatively charged chemicals) and cations (positively charged chemicals) bind together with ionic bonds (electrostatic attraction). Sodium chloride, NaCl (table salt), is an
57
Chemical Bonding Fundamentals example of an ionic compound. A sodium ion, having given up an electron (e-) in an attempt to obtain a filled outer shell, is positively charged since the number of protons exceeds the number of electrons.
Na → Na+ + eChloride, on the other hand, accepts an electron to fill its outermost shell.
Cl + e- → ClThe sodium ion (Na+) is the cation and the chloride ion (Cl-) is the anion. A bond can now form between the negatively-charged Cl- and the positively-charged Na+. The above reaction can be written as:
Na+ + Cl- → Na+ClIn covalent bonds, the electrons can be either shared equally or unequally between atoms. When electrons in a covalent bond are share equally, the resulting molecule is non-polar.
There is not overall charge associated with that molecule. Unequal sharing results in a polar molecule. There exists a positive and negative “pole” or area within polar molecules. Fats are typically non-polar while water is a polar molecule. Polar molecules can readily interact with other polar molecules. The positive area of one polar molecule and the negative area of another polar molecule attract each other in an electrostatic manner. What happens when oil is added to water? Do they Figure 5: Molecular polarity is present in wainteract? Polar and non-polar molecules do not, in fact, inter- ter. The negative region is focused near the oxygen molecule, while the positive region is act. focused near the hydrogen molecules.
Figure 6: Olive oil (a fat) is non-polar, while vinegar (a water-based solution) is polar. These two solutions are immiscible, or, cannot form a homogenous mixture.
58
Chemical Bonding Fundamentals
Pre-lab Questions
1. List the atomic numbers for each of the following elements.
Iron
Oxygen
Calcium
Nitrogen
Potassium
Hydrogen
2. What determines if a bond is polar?
3. Use the periodic table to determine if potassium chloride (KCl) formed through covalent or ionic bonds?
Use evidence from the Introduction to support your answer.
4. Research two common, polar molecules and two common nonpolar molecules. Draw their molecular structure and explain how the structure makes each molecule polar or non-polar.
Experiment 1: Slime Time
Inks can be polar or non-polar. Polar solvents pick up polar inks, while non-polar solvents pick up non-polar inks. In this experiment, you will use inks to identify slime and silly putty as polar or non-polar. You will also use paper chromatography to verify the inks are correctly identified as polar or non-polar.
Materials
(1) 250 mL Beaker
Silly Putty®
5 mL 4% Borax Solution, Na2B4O7·10H2O
Dry Erase Marker
Ruler
(1) 10 mL Graduated Cylinder
Uni-ball® Roller Pen
(1) 100 mL Graduated Cylinder
*Distilled or Tap Water
Filter Paper (Disk)
*Newspaper
Filter Paper (Square)
*Notebook Paper
0.5 g Guar Gum
*Scissors
Wooden Stir Stick
Highlighter
Permanent Marker
*You Must Provide
1 Popsicle Stick
59
Chemical Bonding Fundamentals
Procedure
Part 1: Making Slime
1. Weigh out 0.5 g of guar gum into a 250 mL beaker.
2. Measure 50.0 mL of distilled water into a 100 mL graduated cylinder and pour it into the 250 mL beaker that contains the guar gum.
3. Rapidly stir the mixture with a wooden stir stick for three minutes, or until the guar gum is dissolved.
4. Measure 4.00 mL of a 4% Borax solution into a 10 mL graduated cylinder and add it to the guar gum and water. 5. Stir the solution until it becomes slime. This will take a few minutes. If the slime remains too runny, add an additional 1.0 mL of the 4.0% Borax solution and continue to stir until the slime is the slightly runny or gooey. 6. Once you are satisfied with the slime, pour it into your hands. Be sure not to drop any of it on to the floor.
7. Manipulate the slime in your hands. Write down observations made about how slime pours, stretches, breaks, etc. in Part 1 of the Data section.
CAUTION: Slime is slippery and, if dropped, it can make the work area slick.
8. Place the slime back into the beaker and WASH YOUR HANDS.
Part 2: Slime and Putty Ink Tests
1. On a piece of notebook paper make one 20 - 25 mm long mark of each of the inks you are testing
(permanent marker, highlighter, Dry Erase, and Uni-ball® Roller Pen). Space the marks at least one inch apart. Use a pencil to label each mark with its description.
a. Water soluble inks include those in highlighters and certain pens.
b. Water insoluble inks include those in a permanent pen/markers, newsprint, and a dry-erase markers. 2. While the inks are drying, select a passage or a picture in the newspaper to test with the slime.
3. Develop a hypothesis stating whether or not you believe the slime produced in Part 1 will pick up newsprint ink. Record this hypothesis in the Post-Lab Questions section. Then, break off a small piece of slime that is 3 - 5 cm in diameter. Gently place this piece on top of the newspaper print, then carefully pick it up again.
4. Observe and record in Table 1 whether or not the ink was picked up onto the slime.
60
Chemical Bonding Fundamentals
5. Break off another small piece of slime. Once the inks from Step 1 have dried gently place the slime on top of the first spot on the notebook paper, then carefully pick it up. Repeat this for each of the inks. Observe and record which inks were picked up (dissolved) by the slime in Table 1.
6. Repeat this ink testing two more times for accuracy.
7. Hypothesize which inks the silly putty will pick up in the Part 2 of the Data section. Then, perform the ink tests with the Silly Putty® according to the procedure outlined in Steps 5 - 6.
Part 3: Chromatography of Ink Samples
1. Use a pencil or scissors to poke a small hole in the center of a piece of filter paper (see Figure 7).
Folded wick
2. Spot the filter paper evenly spaced approximately 2 cm from the small hole with the two insoluble inks and the
Ink spots
two soluble inks that were used in Part 2, Step 1.
3. Obtain a ½ piece of filter paper. Fold the paper in half several times so that it makes a narrow wick.
4. Insert the wick into the hole of the spotted paper so that it is above the top of the filter paper by approximately 2 cm.
5. Fill a 250 mL beaker 3/4 full with water.
6. Set the filter paper on top of the beaker so that the bottom of the wick is in the water. The paper should hang over the edge of the beaker with the spotted side up.
7. Allow water to travel until it is approximately 1 cm from
Figure 7: Chromatography apparatus for
Procedure Part 3.
the edge of the filter paper. Remove the filter paper from the beaker.
8. Observe which inks moved from where they were originally spotted. Record your observations in Part 3 of the Data section.
61
Chemical Bonding Fundamentals
Table 1: Results of Ink Testing for Silly Putty®
Picked up (dissolved)
Name of Ink
Trial 1
Trial 2
Trial 3
Newsprint
Highlighter
Uni-ball® Roller Pen
Permanent Marker
Dry Erase Marker
Data
Part 1
•
Slime Observations:
Part 2
•
Hypothesis for Silly Putty ® (Procedure Part 2, Step 7):
Part 3
• Observations of inks following chromatography:
62
Did not pick up
Trial 1
Trial 2
Trial 3
Chemical Bonding Fundamentals
Post-Lab Questions
1. Record your hypothesis regarding the slime’s ability to pick up newsprint ink here.
2. Did the slime pick up water soluble or water insoluble inks? From these results, what can you conclude about the polarity of slime molecules?
3. Explain how you determined your hypothesis about whether or not silly putty would pick up water soluble inks. Was your hypothesis correct?
4. Were the inks you used properly classified as soluble and insoluble? Explain your answer.
63
Lab 4
Introduction to the Microscope
Introduction to the Microscope
Learning Objectives
•
Demonstrate and explain how to use a microscope
•
Explain how different types of microscopes affect magnification and resolving power
•
Identify parts of the a microscope
Introduction
Microbiology is the study of microorganisms. Microorganisms are organisms that are too small to be seen with the human eye. This is a huge field for such small subjects.
There are many times more microorganisms on Earth than all the other organisms put together. It is estimated, for example, that in the human body alone there are 10 microorganisms for every one human cell. That means that 90% of the cells found in the human body are not human! There are many other objects besides microorganisms, such as biological structures, that cannot be seen with the human eye. What type of objects can you think of that cannot be seen without the aid of some magnification? Biological structures might include a valve found in a vein that prevents blood from moving backwards or the cells that make up a multicellular organism. The fact is Figure 1: Bacterial species exhibit differing shapes. that there are an unimaginable number of objects that we cannot see with the naked eye.
Types of Microscopes
Microscopes are used for the visualization of objects too small for us to see. There are many types of microscopes that range from low-level magnification (e.g., hand-held magnification lens) to mid-level magnification
(e.g., compound light microscopes) to very high-power magnification (e.g., an electron microscope). The type of microscope that is used depends on the application. For example, if you were trying to locate and remove a small splinter from your finger, a hand-help magnification lens (magnifying glass) would be extremely helpful. If you wanted to examine the structure of a tiny virus, you would need an electron microscope.
67
Introduction to the Microscope
Magnification
Magnification is one characteristic to consider when deciding what type of microscope is needed for a particular application. The other key characteristic is the resolving power. The resolving power refers to the ability of a microscope to show detail. Having a device that can greatly magnify an object is helpful only if the magnified object is well defined (not fuzzy). The resolution is largely dependent on the wavelength of light used. Shorter wavelengths produce higher resolutions. Light microscopes use visible light and electron microscopes use a beam of electrons that travel in wavelike patterns. These waves are 100,000 times shorter than visible light waves! Electron microscopy can therefore be used to visualize extremely small structures.
The compound light microscope is the most commonly used type of microscope. The name compound refers to presence of more than one lens. The use of these microscopes can be traced to the late 1500s. The early compound microscopes could magnify objects up to about 10X (this means 10 times). In the 1600s a
Dutch linen merchant, Antonie van Leeuwenhoek, greatly improved upon the production of lens and was able to create microscopes that could magnify objects 200X. He observed and recorded many types of microorganisms with his microscopes. Because of this, van Leeuwenhoek is frequently referred to as “the Father of
Microbiology”. Modern day compound light microscopes can typically magnify objects up to 1000X.
68
Figure 2: Microscope components.
Introduction to the Microscope
A representative compound light microscope is shown in Figure 2. Note the two sets of lenses:
•
The ocular lenses (close to your eyes)
•
The objective lenses (close to the “object” on the stage)
Along with a light source, these lenses work together to magnify the object being viewed. In the case of the compound light microscope, the total magnification is equal to the magnification power of the ocular lens multiplied by the magnification power of the objective lens. For example, if the ocular lens magnifies 10X and the objective lens magnifies 10X, the total magnification is 100X.
Microscope Components
Below is a list of the parts of a compound light microscope. Refer to Figure 2 as you read through this list to familiarize yourself with these parts.
•
Base: The flat support of the microscope.
•
Light: Illuminates the object being viewed. This can be either in the form of a light source or a mirror that reflects ambient light onto the image. In the latter case it is important to be working in an environment with adequate ambient light.
•
Stage: Supports the slide or other material to be viewed.
•
Diaphragm: Controls the amount of light allowed on the object.
•
Stage Clips: Secure the slide in place.
•
Revolving Nosepiece: Rotates the objective lenses of different magnifications and allows one of them to be positioned over the slide.
•
Arm: Connects the lower base and the upper head of the microscope (also used to carry the microscope). •
Head: Supports both the ocular lens and the revolving nosepiece.
•
Ocular Lens (eyepiece): The lenses on the microscope typically have a magnification of 10X. If your microscope has a pointer, which is used to indicate a specific area of the specimen, it is attached here. Monocular Microscopes have a single ocular eyepiece while binocular microscopes have two ocular eyepieces.
69
Introduction to the Microscope
How to Use a Microscope
The following steps describe the proper use of a compound light microscope.
1. Always carry a microscope with one hand securely around the arm and the other underneath the base for support.
2. Place the microscope on a table, plug it in, and turn on the light source (or adjust the mirror as necessary). Note: When cleaning a microscope, do not use paper towels or cloths as this will scratch the lens. To preserve the microscope, use only lens paper that will not scratch the optics.
3. To prevent damage to the lens or slides, always start and end with the scanning power objective lens (the shortest one) above the light source.
4. Place your slide on the stage and secure it with the stage clips. It is helpful to visually orient the slide so the object to be viewed is directly in the middle of the opening in the stage where the light is directed up toward the slide.
5. Turn the course adjustment knob to bring the stage all the way up to the scanning power objective lens. While looking through the lens, use the course adjustment knob to slowly lower the stage until the specimen comes into focus.
Note: When using a binocular microscope, adjust the distance between the two oculars until only one object is seen. Record this distance and set your microscope to this distance every time you use it. If someone else uses the microscope, the lenses may be re-adjusted for their eyes.
6. To adjust the light, open or close the diaphragm located over the light source. When properly illuminated, the specimen should not be gray or exceptionally bright.
7. With the object is in general focus, rotate the revolving nosepiece to the low-power lens (the next longest). After focusing with the course adjustment knob, switch to the fine adjustment knob to obtain more precise and greater detail. It may also be necessary to adjust the light, because more light reduces contrast (sharpness).
8. To become familiar with the mechanical stage knobs around the base of the microscope (if present), turn one slowly to the right, noting that the image will be moving toward the left. This image inversion is caused by the lenses.
9. If you need higher magnification, slowly rotate the high-power lens into place (the next longest lens). This will bring the tip of the lenses very close to the slide.
10. Make sure the objective lens does not touch the slide.
11. Whenever you use the high-power lens, only use the fine adjustment knob. If the object was well focused while viewing with the low-power lens, very little adjustment should be necessary.
70
Introduction to the Microscope
12. If you cannot bring the object into focus, return to the low-power lens, focus the object, and then return to the high-power lens.
13. When finished, move the revolving nosepiece to the scanning objective lens position before removing the slide.
Specimen Preparation
Proper specimen preparation helps produce the best visualization possible with any microscope. Glass slides are used to contain the specimen. Live specimens are usually prepared as wet mounts. In wet mount preparation, the specimen is placed on the slide and a much thinner glass cover slip is placed on top of the specimen. Samples are frequently heated to simultaneously kill and fix (secure) the sample to the glass slide.
Fixed samples are then stained to enhance the observable contrast between the cellular features.
?
Did You Know...
Different types of stains may be used depending on the desired outcome. For example, if a specific structure is being studied, one might select a dye that stains only that unique structure. Perhaps the most well-known stain is the Gram stain. The Gram stain was developed in
1884 by Hans Christian Gram. Gram staining is often used to categorize bacteria as
Gram positive or Gram negative. The
Gram stain distinguishes between bacteria with thick cell walls (Gram positive Gram negaƟve bacterial populaƟon. Image courtesy of species) from those with thin cell walls the CDC Image Library.
(Gram negative species). Thick cell walls entrap the crystal violet, resulting in purple colored cells. Thin cell walls do not entrap the crystal violet dye. Instead, the safranin stain is accepted and red colored cells result.
How to Prepare a Wet Mount Slide
1. To make a wet mount for a specimen that is not already in liquid, take a clean slide and place the specimen in the center.
2. Add one drop of water.
Note: For cells that are transparent, it may be necessary to add a small drop of stain as opposed to water.
3. Carefully add a coverslip by placing one end down and slowly lowering the other end.
71
Introduction to the Microscope
Note: If the coverslip is added too quickly, large air bubbles may become trapped which can cause difficulty viewing the slide. If this happens, gently remove the coverslip, add another drop of water and try again.
4. Remove excess liquid on the bottom of the slide or around the edges before it is placed on the microscope to avoid damage to the lens. Just touch a tissue to the edge of the coverslip to draw away the water (this is an example of capillary action).
5. If the specimen is already in liquid, place a drop in the middle of the slide and add the coverslip as you did in Step 3.
Pre-Lab Questions
1. Label the following microscope using at least four of the components described within the Introduction.
72
Introduction to the Microscope
Experiment 1: Virtual Magnification Exercise
Materials
“How Big Is It?” demonstration on Student Portal
Note: Review the directions for signing in to the Student Portal at the beginning of this manual if uncertain how to access this information
Procedure
1. Log into your eScience Student Portal account (www.eScienceLabs.com/portal/) and locate the “How Big
Is It” demonstration located near the end of the Introduction to the Microscope section.
2. Load the animation and beginning with the head of a pin, increase the magnification by clicking the arrows below the picture. Note the relative sizes of the objects on the pinhead.
3. Be sure to notice the magnification bar on the lower portion of the demonstration that shows the magnification required to see the objects.
Post-Lab Questions
1. At what magnification do you first notice the ragweed pollen?
2. Which is bigger, a rhinovirus or E. Coli?
3. Based on the magnification, how many of the E. Coli can fit into the same space as the head of a pin?
4. About how many red blood cells could fit across the diameter of a human hair (again, look at the magnification scale)?
73
Introduction to the Microscope Experiment 2: Virtual Microscope
Materials
Virtual Microscope on Student Portal
Note: Review the directions for signing in to the Student Portal at the beginning of this manual if uncertain how to access this information.
Procedure
1. Log into your eScience Labs Student Portal account and locate the Virtual Microscope activity located near the end of the Introduction to the Microscope section.
2. Take a tour of the virtual microscope by clicking the “Start Tour” button on the right hand side and learn how to use the different controls to effectively use the simulation.
3. Once you are comfortable using the virtual microscope, turn the light on. Switch views so that you are looking through the ocular lens and can look at the slides.
4. Select the letter e slide on the top right of the page and examine with the 4X objective lens. Increase the magnification until the letter no longer fits in the field of view. Note this magnification.
5. Select the cheek smear slide on the right side of the page and bring the cells within the red circle into focus using the 4X, then 10X objective lenses.
6. View the slide under the 40X and 100X objective lenses, making sure you stay within the red circle.
7. Next, select the onion root tip slide on the right side of the page to view. Start with the 4X objective lens, and bring the cells within the red circle into focus.
8. Switch to the 10X objective lens and readjust the focus so the slide is clear. Continue looking at the slide under higher magnification using the 40X and 100X objective lenses.
9. On the left hand side of the screen, select the “Try This” box. Under measurement, select the m1 box to open an activity that will instruct you how to measure the letter e. (Remember this number as you will have to report it in Question 5!)
Note: You may have to go through the tutorial tour again in order for the “Try This” box to become available. Be sure to work your work through the entire checklist, and then click “exit” button to prompt the “Try This” button to appear again.
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Introduction to the Microscope Post-Lab Questions
1. After completing the m1 exercise in the “Try this” section, how tall is the letter e?
2. What is the highest objective lens you can use to see the entire letter e?
3. The nuclei (the structure inside a cell that contains DNA) of the cheek cells have been stained using a special dye so that they appear purple. What shape are they?
4. At high magnification, you may notice that not all of the nuclei in the onion root tip slide appear as the shape you described in the question above. What do they look like?
5. What is the first step normally taken when you look through the ocular lenses?
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Lab 5
The Chemistry of Life
The Chemistry of Life
Learning Objectives
•
Compare and contrast organic and inorganic molecules
•
Relate hydrogen bonding to macromolecules found in living things
•
Compare and contrast the four major organic macromolecules: lipids, carbohydrates, nucleic acids and proteins
Introduction
There are over 100 known elements in the periodic table. Elements are pure substances made of only one type of atom. Interestingly, the number of naturally occurring elements is unclear. More than 90% of all matter is composed of combinations of just four of the approximately 88 naturally occurring elements: oxygen, carbon, hydrogen, and nitrogen. Living organisms, in general, require about 20 elements naturally occurring elements. The periodic table continues to develop as new elements are synthesized in laboratories.
Figure 1: The periodic table of elements categorizes all of the known elements. Groups are listed vertically as 1 7. Periods are listed horizontally as 1 - 18.
79
The Chemistry of Life
Organic Molecules
Although the term “organic” is frequently used to refer to foods or clothing, in biology and chemistry, the terms organic and inorganic are used to categorize molecules by their origin. All organic molecules are of biological origin. In contrast, inorganic molecules are not of derived from living organisms. Since organic molecules are those that derived from living organisms, it follows that they could not be produced without life. The human body is made up of roughly 27% organic molecules and 73% inorganic molecules. Carbohydrates (sugars) and proteins are examples of organic molecules (note that both of these types of molecules are produced by living organisms).
Figure 2: Glucose, fructose, and galactose are all examples of carbohydrates.
Carbon dioxide (CO2), table salt (NaCl) and water (H2O) are all examples of inorganic molecules. Notice that carbon dioxide (CO2) contains carbon but is not an organic molecule. If a molecule contains carbon it may be organic, but this is not the only requirement. Organic molecules are typically identified by the presence of carbon-hydrogen bonds.
Macromolecules
There are many classes of organic compounds, based on the functional groups they contain. In living organisms, the most important organic com- Figure 3: Carbon dioxide is a pounds belong to a classification of molecules called macromolecules. major gas exhaled by humans.
The term “macromolecule” simply means a large molecule. Recall that mol80
The Chemistry of Life ecules are substances that contain two or more atoms bonded together. The same four types of macromolecules are used by all living organisms for cellular metabolism and reproduction. These common biological macromolecules are lipids, nucleic acids, carbohydrates and protein. The properties they convey are of great importance to cell function.
Lipids, or fats, have many functions within living organisms including energy storage, membrane structure, and aids in the formation of internal cellular components. All genetic material is composed of nucleic acids (DNA). Another nucleic acid, RNA, functions in the production of proteins and other cellular processes. Carbohydrates are commonly referred to as sugars. This class of organic compounds serves as one of the primary sources of metabolic energy.
The carbohydrate monosaccharide subunit most commonly used for energy is glucose. Other useful carbohydrates include maltose, lactose, sucrose, and starch. Proteins are the most abundant macromolecule in living systems. Like lipids, proteins perform a variety of functions. For example, proteins are major
Figure 4: Omega-3 Fatty acid. components of tendons, ligaments and muscles in the human body.
Chemical Bonding
Macromolecules are formed by the covalent bonding between subunits. Recall that covalent bonds can produce molecules that are either polar or non-polar. Lipids, for the most part, are non-polar. Some of the 20 amino acids found in proteins are non-polar and some are polar. When macromolecules are created, non-polar regions tend to be located close together. Polar regions can participate in another type of chemical bond called hydrogen bonding. Hydrogen bonds result from the interaction of the positive region of one polar molecule and the negative region of another polar molecule.
Although relatively weak bonds, these are extremely imFigure 5: Approximately 70% of a human adult portant in the maintenance of chemical structures. In fact, the body is composed of water. double strands found in DNA as well water molecules are held together by hydrogen bonds. Adenine and thymine
(nucleic acids found in DNA) are held together via two hydrogen bonds, and cytosine and guanine are held together by three.
Living things require a constant supply of energy. Throughout this manual, you will learn about the reactions that take place inside of organisms. The sum of these reactions is called metabolism, and is a general term used to describe the energy required to keep those reactions occurring.
81
The Chemistry of Life
Pre-Lab Questions
1.
Nitrogen fixation is a natural process by which inert or unreactive forms of nitrogen are transformed into usable nitrogen. Why is this process important to life?
2. Given what you have learned about the hydrogen bonding shared between nucleic acids in DNA, which pair is more stable under increasing heat: adenine and thymine, or cytosine and guanine? Explain why.
3. Which of the following is not an organic molecule; methane (CH4), fructose (C6H12O6), rosane (C20H36), or ammonia (NH3)? How do you know?
Experiment 1: Testing for Proteins
The protein molecules in many foods provide the amino acid building blocks required by our own cells to produce new proteins. To determine whether a sample contains protein, a reagent called Biuret solution is used. Biuret solution contains copper ions. However, the chemical state of the copper ions in Biuret solution causes them to form a chemical complex with the peptide bonds between amino acids (when present), changing the color of the solution. Biuret solution is normally blue, but changes to pink when short peptides are present and to violet when long polypeptides are present.
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Figure 6: Biuret solution only is located on the far left side of the image (blue).
Note the transition from blue to violet as proteins are added to the solution, causing the solution to transition from blue to violet.
The Chemistry of Life
Materials
(2) 250 mL Beakers
5 Test Tubes (Plastic)
25 Drops Biuret Solution, H2NC(O)NHC(O)NH
Test Tube Rack
(1) Knox® Gelatin Packet
5 mL Unknown Solution
5 mL 1% Glucose Solution, C6H12O6
*Tap Water
(1) 10 mL Graduated Cylinder
*Hot Water
(1) 100 mL Graduated Cylinder
*Egg White
Permanent Marker
5 Pipettes
*You Must Provide
Procedure
1. Label five test tubes 1, 2, 3, 4 and 5.
2. Prepare your testing samples as follows:
a. Mix one egg white with 25 mL water in a 250 mL beaker to create an albumin solution. Pipette
5 mL of this solution into Test Tube 1.
b. Mix the packet of Knox® gelatin with 50 mL hot water in a second 250 mL beaker. Stir until dissolved. Pipette 5 mL of this solution into Test Tube 2.
3. Pipette 5 mL of the 1% glucose solution into Test Tube 3.
4. Use the 10 mL graduated cylinder to measure and pour 5 mL of water into Test Tube 4.
5. Pipette 5 mL of the “Unknown Solution” into Test Tube 5.
6. Record the initial color of each sample in Table 1.
7. Develop a hypothesis regarding what you predict will happen when Biuret solution is added to Tubes 1
- 4. Record your hypothesis in the Post-Lab Question section. Then, pipette five drops of Biuret solution to each test tube (1 - 5). Swirl each tube to mix.
8. Record the final color in Table 1.
Note: Protein is present in the sample if a light purple color is observed.
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The Chemistry of Life
Table 1: Testing for Proteins Results
Sample
Initial Color
Final Color
Protein Present
1 - Albumin Solution
2 - Gelatin Solution
3 - Glucose
4 - Water
5 - Unknown
Post-Lab Questions
1. Record your hypothesis about what will happen when Biuret solution is mixed with the solutions from test tubes 1, 2, 3, and 4 here. Be sure to use scientific reasoning to support your hypothesis.
2. Write a statement to explain the molecular composition of the unknown solution based on the results obtained during testing with the Biuret solution and each sample solution.
3. Diet and nutrition are closely linked to the study of biomolecules. How should you monitor your food intake to insure the cells in your body have the materials necessary to function?
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The Chemistry of Life
4. There are other types of reagents used to determine what type of biomolecule a substance is. For example, copper ions present in Benedict’s reagent reacts with the free end of any reducing sugars, such as glucose, when heated. Originally blue in color, these copper ions are reduced by the sugar, and produce an orange-red colored precipitate. Alternatively, iodine-potassium iodide (IKI) may also be used when working with starch. IKI contains special tri-iodine ions which interact with the coiled structure of a starch polymer. Prior to a reaction, the IKI displays a yellow-brown color; however, after reacting with starch, a dark purple or black color is presented.
The molecule pictured below produced a blue color when tested with Benedict’s reagent, a yellow color when tested with IKI, and a violet color when tested with Biuret reagent. Based on the structure shown below and these chemical results, what kind of biomolecule is this?
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Lab 6
Diffusion
Diffusion
Learning Objectives
•
Apply diffusion to cellular activity and dialysis
•
Explain how solute size, molecular polarity and membrane permeability affect the rate of diffusion
•
Explain how concentration gradients and molarity affect the direction of diffusion
Introduction
Molecules are constantly in motion due to the kinetic energy present in every atom. This energy results in the net movement of molecules from areas of high concentration to areas of low concentration, a process called diffusion. If uninhibited, this net movement will continue until equilibrium is reached and the molecules are uniformly distributed. Once equilibrium is achieved, molecules will continue to move in each direction at an equal rate.
Diffusion
A solution contains two or more substances (solutes) that have been dissolved by a solvent. In the context of a cell, the intracellular and extracellular fluids are the solvents which contain dissolved material (solutes).
The rate of diffusion depends on membrane characteristics, size of the solute, and molecular polarity. Because the medium will not change in a biological system, the diffusion rate is usually dictated by molecular characteristics of the solute. Small, non-polar molecules exhibit a higher rate of diffusion than large, charged ones.
Diffusion Factors
The direction of diffusion depends on concentration gradients, heat, and pressure. The concentration gradient is the change of molecular density over a given area (density is defined as unit mass per volume of space). Molarity (moles/ liter) is a way to express concentration. It is the number of moles of solute dissolved in one liter of solution, and is commonly represented by ‘M’. A mole equals 6.02 x 1023 molecules or atoms. Molecular weight (MW) is the weight of one mole of a chemical and is based off the atomic mass of each atom in the chemical formula. Once the molecular weight of a chemical is known, the weight of chemical to dissolve in a solvent for a specific molar solution can easily be calculated.
Figure 1: Molecules diffuse down concentra-
Temperature and pressure typically remain constant in biologtion gradients to create equilibrium. ical systems, making the concentration gradient the best indi-
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Diffusion cator of directionality. In general, molecules will move towards areas of lower concentration.
Diffusion Across a Membrane
A major determinant of diffusion within a biological system is membrane permeability. Cells, as well as some organelles within the cell, are surrounded by selective and differentially permeable membranes. These membranes control the interaction of the cell and its surrounding environment. Acting as a living gatekeeper, the membrane allows, slows, or denies access into the cell.
Cellular membranes are composed of two layers of phospholipids (Figure 1). Each phospholipid has a hydrophobic, fatty acid tail and a hydrophilic head. This lipid bilayer selects for molecules that can dissolve into the lipid environment and against those that cannot. The ability of a molecule to cross the membrane is also determined by its size and electrical charge. Figure 2: Diffusion through a phospholipid
Small, uncharged molecules often pass through the membrane bilayer (cell membrane). easily, while most large or charged molecules are prevented from passing. Molecules which cannot diffuse across the membrane may be able to cross through other regulated gateways located within the membrane.
Dialysis is the separation of molecules through diffusion. In dialysis, a differentially permeable membrane is used to separate the components of a mixed solution containing more than one type of molecule. This membrane allows the free passage of water, but limits the movement of molecules by their size. In one of the following experiments, you will dialyze a solution of glucose and starch to observe membrane permeability.
?
Did You Know...
Serial Dilutions
The concentration of a solution may also impact the membrane permeability. A solution or its components may not be able to pass through a membrane if the concentration is too high. For this reason, serial dilutions may be performed to create solutions of lower concentrations. The original sample is a known volume and concentration. A small volume of this sample is transferred sequentially among new tubes, called dilution blanks. The sample is mixed with water (or another diluent) throughout the process to create more dilute solutions (Figure 3).
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Hemodialysis is a method of removing toxic substances from the blood when the kidneys are unable to do so. It is frequently used for patients with kidney failure, but may also be used to quickly remove drugs or poisons in dangerous situations.
Diffusion
9 mL
9 mL
9 mL
9 mL
9 mL
9 mL
Figure 3: Serial dilution schematic. The dilution blanks all have 9 mL of diluent in them such that
1
when 1 mL is added, it represents /10th the concentration of the previous tube. The dilution of the sample is written below each tube.
Pre-Lab Questions
1. A concentration gradient affects the direction that solutes diffuse. Describe how molecules move with respect to the concentration.
2. How does the size of a solute affect the rate of diffusion? Consider the size and shape of a molecule in your response.
3. Does polarity affect diffusion? Explain your answer using scientific principles.
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Diffusion
Experiment 1: Diffusion through a Liquid
In this experiment, you will observe the effect that different molecular weights have on the ability of dye to travel through a viscous medium.
Materials
1 60 mL Corn Syrup Bottle, C12H22O11
Ruler
Red and Blue Dye Solutions (Blue molecular weight = 793 g/mole; Red molecular weight = 496 g/mole) *Stopwatch
(1) 9 cm Petri Dish (top & bottom halves)
*Tape
*You Must Provide
Procedure
1. Use clear tape to secure one half (either the bottom or the top half is fine) of the petri dish over a ruler.
Make sure that you can read the measurement markings on the ruler through the petri dish. The dish should be positioned with the open end of the dish facing upwards.
2. Carefully fill the half of the petri dish with corn syrup until the entire surface is covered.
3. Develop a hypothesis discussing which dye you believe will diffuse faster across the corn syrup and why.
Record this in the Post-Lab Questions section. Then, place a single drop of blue dye in the middle of the corn syrup. Note the position where the dye fell by reading the location of the outside edge of the dye on ruler. 4. Record the location outside edge of the dye (the distance it has traveled) every ten seconds for a total of two minutes. Record your data in Tables 1 and 2.
5. Repeat Steps 1 - 4 using the red dye, the second half of the petri dish, and fresh corn syrup.
Table 1: Rate of Diffusion in Corn Syrup
Time (sec)
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Blue Dye
Red Dye
Time (sec)
10
70
20
80
30
90
40
100
50
110
60
120
Blue Dye
Red Dye
Diffusion
Table 2: Speed of Diffusion of Different Molecular Weight Dyes
Structure
Molecular Weight
Total Distance
Traveled (mm)
Speed of Diffusion
(mm/hr)*
Blue Dye
Red Dye
*Multiply the total distance diffused by 30 to get the hourly diffusion rate
Post-Lab Questions
1. Record your hypothesis from Step 3 here. Be sure to validate your predictions with scientific reasoning.
2. Which dye diffused the fastest?
3. Does the rate of diffusion correspond with the molecular weight of the dye?
4. Does the rate of diffusion change over time? Why or why not?
5. Examine the graph below. Does it match the data you recorded in Table 2? Explain why, or why not.
Submit your own plot if necessary.
Diffusion (mm)
Speed of Diffusion of Different Molecular Weight Dyes
Time (seconds)
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Diffusion
Experiment 2: Concentration Gradients and Membrane Permeability
In this experiment, you will dialyze a solution of glucose and starch to observe:
•
The directional movement of glucose and starch.
•
The effect of a selectively permeable membrane on the diffusion of these molecules.
An indicator is a substance that changes color when in the presence of the substance it indicates. In this experiment, IKI will be used an indicator to test for the presence of starch and glucose.
Materials
(5) 100 mL Beakers
*Stopwatch
10 mL 1% Glucose Solution, C6H12O6
*Water
4 Glucose Test Strips
*Scissors
(1) 100 mL Graduated Cylinder
*15.0 cm Dialysis Tubing
4 mL 1% Iodine-Potassium Iodide, IKI
5 mL Liquid Starch, C6H10O5
*You Must Provide
3 Pipettes
*Be sure to measure and cut only the length you
4 Rubber Bands (Small; contain latex, handle with
for later experiments.
gloves on if allergic)
Attention!
Do not allow the open end of the dialysis tubing to fall into the beaker. If it does, remove the tube and rinse thoroughly with water before refilling with a starch/glucose solution and replacing it in the beaker.
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need for this experiment. Reserve the remainder
Note:
•
Dialysis tubing can be rinsed and used again if you make a mistake. •
Dialysis tubing must be soaked in water before you will be able to open it up to create the dialysis “bag”. Follow the directions for the experiment, beginning with soaking the tubing in a beaker of water. Then, place the dialysis tubing between your thumb and forefinger and rub the two digits together in a shearing manner. This should open up the
"tube" so you can fill it with the different solutions.
Diffusion
Procedure
1. Measure and pour 50 mL of water into a 100 mL beaker. Cut a piece of dialysis tubing 15.0 cm long.
Submerge the dialysis tubing in the water for at least 10 minutes.
2. Measure and pour 82 mL water into a second 100 mL beaker. This is the beaker you will put the filled dialysis bag into in Step 9.
3. While the dialysis bag is still soaking, make the glucose/sucrose mixture. Use a graduated pipette to add five mL of glucose solution to a third beaker and label it “Dialysis bag solution”. Use a different graduated pipette to add five mL of starch solution to the same beaker. Mix by pipetting the solution up and down the pipette six times.
4. Using the same pipette that you used to mix the dialysis bag solution, remove two mL of that solution and place it in a clean beaker. This sample will serve as your positive control for glucose and starch.
a. Dip one of the glucose test strips into the two mL of glucose/starch solution in the third beaker.
After one minute has passed, record the final color of the glucose test strip in Table 3. This is your positive control for glucose.
b. Use a pipette to transfer approximately 0.5 mL of IKI to into the two mL of glucose/starch solution in the third beaker. After one minute has passed, record the final color of the glucose/starch solution in the beaker in Table 3. This is your positive control for starch.
5. Using a clean pipette, remove two mL of water from the 82 mL of water you placed in a beaker in Step 2 and place it in a clean beaker. This sample will serve as your negative control for glucose and starch.
a. Dip one of the glucose test strips into the two mL of water in the beaker. After one minute has passed, record the final color of the glucose test strip in Table 3. This is your negative control for glucose. b. Use a pipette to transfer approximately 0.5 mL of IKI to into the two mL of water in the beaker.
After one minute has passed, record the final color of the water in the beaker in Table 3. This is your negative control for starch.
Note: The color results of these controls determine the indicator reagent key. You must use these results to interpret the rest of your results.
6. After at least 10 minutes have passed, remove the dialysis tube and close one end by folding over 3.0 cm of one end (bottom). Fold it again and secure with a rubber band (use two rubber bands if necessary).
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Diffusion
7. Make sure the closed end will not allow a solution to leak out. You can test this by drying off the outside of the dialysis bag with a cloth or paper towel, adding a small amount of water to the bag, and examining the rubber band seal for leakage. Be sure to remove the water from the inside of the bag before continuing.
8. Using the same pipette which was used to mix the solution in Step 3, transfer eight mL of the solution from the Dialysis Bag Solution beaker to the prepared dialysis bag.
9. Place the filled dialysis tube in beaker filled with 80 mL of water with the open end draped over the edge of the beaker as shown in Figure 4.
10. Allow the solution to sit for 60 minutes. Clean and dry all materials except the beaker with the dialysis bag.
11. After the solution has diffused for 60 minutes, remove the dialysis tube from the beaker and empty the contents into a clean, dry beaker. Label it dialysis bag solution.
Figure 4: Step 9 reference.
12. Test the dialysis bag solution for the presence of glucose and starch. Test for the presence of glucose by dipping one glucose test strip into the dialysis bag directly. Again, wait one minute before reading the results of the test strips. Record your results for the presence of glucose and starch in Table 4. Test for the presence of starch by adding two mL IKI. Record the final color in Table 4 after one minute has passed.
13. Test the solution in the beaker for glucose and starch. Use a pipette to transfer eight mL of the solution in the beaker to a clean beaker. Test for the presence of glucose by dipping one glucose test strip into the beaker. Wait one minute before reading the results of the test strip and record the results in Table 4. Add two mL of IKI to the beaker water and record the final color of the beaker solution in Table 4.
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Diffusion
Table 3: Indicator Reagent Data
Indicator
Starch Positive
Starch Negative
Glucose Positive
Glucose Negative
Control (Color)
Control (Color)
Control (Color)
Control (Color)
n/a
n/a
IKI Solution
Glucose Test Strip
n/a
n/a
Table 4: Diffusion of Starch and Glucose Over Time
Indicator
Dialysis Bag After 1 Hour
Beaker Water After 1 Hour
IKI Solution
Glucose Test Strip
Post-Lab Questions
1. Why is it necessary to have positive and negative controls in this experiment?
2. Draw a diagram of the experimental set-up. Use arrows to depict the movement of each substance in the dialysis bag and the beaker.
3. Which substance(s) crossed the dialysis membrane? Support your response with data-based evidence.
4. Which molecules remained inside of the dialysis bag?
5. Did all of the molecules diffuse out of the bag into the beaker? Why or why not?
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Lab 7
Osmosis
Osmosis
Learning Objectives
•
Explain how tonicity affects the direction of osmosis
•
Hypothesize why aquaporins are necessary to support osmosis in living things
•
Relate osmotic pressure to water potential
Introduction
The movement of water across a selectively permeable membrane, like the plasma membrane of the cell, is called osmosis. Osmosis is directed from an area of high water concentration to an area of low water concentration. Ultimately, membrane selectivity and the movement of water in and out of the cell regulate the concentration of intracellular material. As solute concentration increases, solvent concentration decreases.
In order to achieve equilibrium, the solvent from a low solute concentration solution will move into an area of high solute concentration to create an equal ratio of solvent to solute in both areas. Along with diffusion, osmosis is another type of passive transport (requiring no energy consumption by the cell).
Tonicity is a relative term that describes the solute difference between solutions and determines the net direction of movement of water molecules (osmosis). There are three types of tonicity:
1. Hypertonic: A solution with a higher solute concentration than the solute concentration on the opposite side of the permeable membrane.
2. Hypotonic: A solution with a lower solute concentration than the solute concentration (i.e., a solution with a higher percentage of water than solute) on the opposite side of the permeable membrane.
3. Isotonic: A solution with equal solute concentrations on both sides of the permeable membrane
(Figure 1).
Figure 1: Chemical diffusion can create a variety of states for the cell or even an entire organ.
101
Osmosis
Water Potential
In osmosis, water molecules flow from hypotonic solutions to hypertonic solutions, until the solutions become isotonic. This underscores the concept of water potential. Biologists use this term to describe the tendency of water to leave one place in favor of another. Water always moves from an area of higher water potential to an area of lower water potential. The more negative the water potential, the higher the concentration of solutes in the system.
In most biological systems, cells are hypertonic to their environment and extracellular water flows into them.
If placed in pure water, cells will burst (lyse) as a result of the increased pressure on the membrane from the additional water that diffused into the cell.
Osmotic Pressure
Osmotic pressure (the force required to prevent osmosis), also called pressure potential, is the tendency of a solution to gain water across an ideal, partially permeable membrane. It directly correlates with tonicity
(higher tonicity causes an increase in osmotic pressure). Some cells, such as plant cells, have specialized structures that regulate osmotic pressure and prevent lysis. This pressure, as well as solute concentration, can affect the water potential (high osmotic pressure = low water potential). This relationship is demonstrated by the following formula:
Ψ = Ψp + Ψs
In this equation, Ψ represents water potential, Ψp is pressure potential, and Ψs is the solute potential.
Both diffusion and osmosis are imperative to cell survival. They enable cells to maintain equilibrium of solute concentrations across two sides of a permeable membrane. However, the two processes facilitate solute equilibrium in opposite ways.
Aquaporins
For much of scientific history, the abundance of water molecules throughout the body led many scientists to believe that osmosis alone was a sufficient mechanism for water movement across cellular membranes.
However, in 1992 Peter Agre discovered a unique family of proteins, later termed “aquaporins”, which were also involved in water transport.
Aquaporins are a specific form of integral membrane proteins which span lipid bilayers to facilitate molecular movement. They use a combination of electromagnetic and hydrophilic interactions and spatial selectivity to funnel water molecules from one side of a membrane to another. Aquaporins are most populous in tissues
102
Osmosis which require high levels of water transport such as the kidney, salivary glands, (lacrimal) tear glands and other epithelial cells. Can you think of a reason why aquaporins are so important in these tissues? Although aquaporins can be ubiquitous in certain locations, the actual aquaporin present may vary by tissue/organ.
The most common aquaporin in called aquaporin 1 (AQP1). AQP1 is found in the kidney and forms a homotetramer protein across the cellular membranes within this organ. “Homotetramer” means that it is composed of four, identical simpler molecules.
Water molecules are attracted to these pores. However, rather than moving through the central homotetramer central channel, water molecules move through one of the four individual channels formed by the four monomer subunits. These channels are very tiny, with a diameter of only 3 Å. This further increases the specific of aquaporins because many molecules are much larger than this (note: water molecules are only 2.8
Å).
Agre won the Nobel Prize in Chemistry in 2003 for this paradigm-changing discovery. You can learn more about Peter Agre and his discovery here: http://www.nobelprize.org/nobel_prizes/chemistry/laureates/2003/ agre-autobio.html Pre-Lab Questions
1. What is the water potential of an open beaker containing pure water?
2. Why don’t red blood cells swell or shrink in blood?
3. How do osmotic power plants work?
4. Research the structures that protect plant and animal cells from damage resulting from osmotic pressure.
Write a few paragraphs explaining what they are, how they work, and where they are located.
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Osmosis
Experiment 1: Tonicity and the Animal Cell
In this experiment you will explore the effects of osmosis on eggs. An egg is surrounded by a permeable membrane and a hard shell. To observe the effects of osmosis the shell will first need to be removed.
Materials
Acetic Acid (Vinegar)
100 mL Graduated Cylinder
4 Pieces of Aluminum Foil (to cover beakers)
100% Sugar Solution (Corn Syrup)
(4) 100 mL Beakers
Stirring Rod
*Distilled Water
*4 Eggs
*You Must Provide
10 mL Graduated Cylinder
Note: This lab requires at least one day to prep and at least one day for observations. For best results, please use a medium or large sized egg. Jumbo or extra large eggs are too large for the experimental set-up.
Procedure
1. Set out four beakers with one egg in each beaker.
Note: Always wear gloves when handling raw eggs.
2. Cover each egg with acetic acid (vinegar).
3. Cover each beaker with a piece of foil and allow the eggs to sit for 24 hours.
Keep foil for entire experiment. You will need it later in the experiment.
4. You will be placing eggs (cells) in different tonicities and observing the results.
Develop a hypothesis and record it in the data section for this experiment.
5. After 24 hours, check to see if the egg shell is dissolved by gently taking out
Figure 2: An egg that h as each egg and rinsing with a small amount of water. The eggs should have a been soaked in vinegar and rinsed with water. yellow tint to them and you should be able to lightly squish the egg between
Some white shell remains.
your fingers (Figure 2). BE CAREFUL NOT TO BREAK THE EGG.
6. If any of the eggs still have a hard shell after 24 hours allow the eggs to sit for another 12 - 24 hours.
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Osmosis
7. Once the egg shells have been dissolved remove them from the vinegar. HANDLE EGGS CAREFULLY. THEY CAN EASILY BREAK.
8. Rinse out and dry all four beakers.
9. Label one beaker “100 % distilled water”, one “100% corn syrup solution”, one “10% corn syrup/90% water solution” and one “control”.
10. In the beaker labeled “10% corn syrup/90% water solution”, prepare a 10% sugar solution by mixing 45 mL of distilled water with 5 mL of corn syrup. Mix well with a stirring rod. Record the initial volume in Table 1.
11. In the beaker labeled “distilled water”, measure and pour 50 mL of distilled water. Record the volume in
Table 1. The egg should be covered. Add or reduce the amount of water if necessary. Record the initial volume in Table 1.
12. In the beaker labeled “100% corn syrup solution”, measure and pour 50 mL of corn syrup (100% sugar solution). The egg should be covered. Add more corn syrup if necessary. Record the initial volume of the solution in Table 1.
13. In the beaker labeled “control”, do not add any solution. Record the volume in Table 1.
14. Add one egg to each beaker and cover with foil.
15. Let the eggs sit for 24 hours. In the meantime, develop a hypothesis stating how you believe each egg will be affected by the solution in the beaker over the 24 hour period. Record your hypothesis in the
Post-Lab Questions section.
16. After 24 hours have passed, observe the eggs. If you do not see any noticeable difference let the eggs sit for another 12 - 24 hours.
17. After the eggs have rested for 24 - 48 hours in the solutions carefully remove them from the beakers.
Make sure to note which eggs came from each beaker. Be careful to not spill the solutions.
18. Record the final volume of each solution using a graduated cylinder and record your results in Table 1.
19. Observe the eggs. Draw your observations in the in Table 2 of data section.
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Osmosis
Data
Hypothesis:
Table 1: Osmosis Results
Solution Type
Volume (mL) Before Osmosis
Volume (mL) After Osmosis
100% distilled water
100% corn syrup
10% corn syrup/ 90% distilled water control Table 2: Observations of Eggs After Osmosis
100% Distilled Water
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100% Corn Syrup
10% Corn syrup/ 90%
Distilled Water
Control
Osmosis
Post-Lab Questions
1. Record your hypothesis from Step 15 here. Be sure to include scientific reasoning to support your predictions.
2. How do each of the three eggs placed in solution compare to the control egg?
3. For each beaker, identify whether the solution inside was hypotonic, hypertonic, or isotonic in comparison to the control beaker.
4. What was the direction of osmosis in the beaker labeled “100% distilled water”? Did the egg in this beaker burst? 5. Was there a volume in the “control” beaker after 24-48 hours? If so, why do you think this is? If not, why do you think this is?
6. Which solution contained a dehydrated cell? Think about someone having a high sugar diet. Hypothesize how their cells might be affected by osmosis?
7. Osmosis is how excess salts that accumulate in cells are transferred to the blood stream so they can be removed from the body. Explain how this process works in terms of tonicity.
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Osmosis
Experiment 2: Tonicity and the Plant Cell
Plant cells are able to generate osmotic pressure while other cells cannot. This is due to specialized plant structures such as the cell wall which prevent lysis caused by osmosis. By taking advantage of this system, you will be able to look at the effects of tonicity in a biological system.
Materials
(1) 100 mL Graduated Cylinder
*Kitchen Knife
Permanent Marker
*2 Potatoes (these must be different types; e.g.,
2 Pipettes
Ruler
16 g Sodium Chloride (Salt), NaCl
russet, Idaho, sweet, etc.)
*Stopwatch
*Water
4 Test Tubes (Glass)
Test Tube Rack
*You Must Provide
*Cutting Board
Procedure
1. Use the permanent marker to label two test tubes as A, and two test tubes as B. Place the test tubes in the test tube rack.
2. Identify the two potato types in the first two cells of the first column in Table 3. Then, select one potato to test first and record observations about the physical characteristics in Table 3.
Note: Be sure to include observations which acknowledge the texture, color, and any other distinguishing factors.
3. Use a knife to carefully cut two strips of the potato on a cutting board. These will be referred to as Sample A and Sample B. The strips should be as close to 10.0 cm. long and 1.0 cm. wide as possible to ensure that the strips fit in the test tubes.
4. Fill the 100 mL graduated cylinder with 50 mL of water. Place the first potato strip (Sample A) into the graduated cylinder and record the initial displacement in Table 3.
Note: Displacement is a measurement of change. It is calculated by subtracting the original volume
(50 mL) from the final volume after the potato is added to the 50 mL of water. For example, 57 mL 50 mL = 7 mL of displacement.
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Osmosis
5. Remove Sample A from the graduated cylinder and place it in Test Tube A. If any water was lost in the graduated cylinder, refill it to the 50 mL graduation mark.
6. Place the second potato strip (Sample B) into the graduated cylinder. Record the initial displacement in
Table 3.
7. Remove Sample B from the graduated cylinder and place it in Test Tube B. If any water was lost in the graduated cylinder, refill it to the 50 mL graduation mark.
8. Repeat Steps 2 - 7 for the second potato type, using the remaining test tubes in the test tube rack.
9. Use a pipette to add water to each of the test tubes with the A samples in them until the water covers the potato strips.
10. Refer to the instructions provided on the bottle with 16 g of sodium chloride in it to create a 20% sodium chloride solution. Use a pipette to add the 20% sodium chloride solution to each of the test tubes with the
B potato samples in them until the solution covers the potato strips.
11. After an hour, pour out the liquid from the test tubes.
12. Repeat Steps 4 - 7 for each sample and type, and record the final displacement values in Table 3.
13. Complete the last column of Table 3 by subtracting the initial displacement from the final displacement.
Table 3: Water Displacement per Potato Sample
Potato
Type
Potato
Sample
Observations
Initial Displacement
(mL)
Final Displacement
(mL) - Step 12
Net Displacement
(mL)
A
B
A
B
Post-Lab Questions
1. How did the physical characteristics of the potato vary before and after the experiment? Did it vary by potato type?
2. What does the net change in the potato sample indicate?
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Osmosis
3. Different types of potatoes have varying natural sugar concentrations. Explain how this may influence the water potential of each type of potato.
4. Based on the data from this experiment, hypothesize which potato has the highest natural sugar concentration. Explain your reasoning.
5. Did water flow in or out of the plant cells (potato cells) in each of the samples examined? How do you know this?
6. Would this experiment work with other plant cells? What about with animal cells? Why or why not?
7. From what you know of tonicity, what can you say about the plant cells and the solutions in the test tubes? 8. What do your results show about the concentration of the cytoplasm in the potato cells at the start of the experiment? 9. If the potato is allowed to dehydrate by sitting in open air, would the potato cells be more likely to absorb more or less water? Explain.
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Lab 8
Enzymes
Enzymes
Learning Objectives
•
Describe enzyme structure and function including activation site
•
Explain how enzymes affect activation energy of a reaction
•
Relate reaction kinetics, activators and inhibitors to the function of enzymes
•
Hypothesize how climate-based changes can affect enzyme function
Introduction
Enzymes are specialized proteins made by living cells that serve as biological catalysts. A catalyst serves to decrease the activation energy normally needed for a reaction to occur naturally. Enzymes can increase the reaction rate between two molecules by over a million times faster than it would be without the enzyme.
Most biochemical reactions require enzymes for them to occur at fast enough rates to be useful. Typical nomenclature for enzymes uses name of the substrate or the chemical reaction it catalyzes as the prefix, and adds the segment “-ase” as the suffix (e.g., catalase, amylase, polymerase, etc.). Keep in mind that some enzymes, such as pepsin, do not follow this nomenclature.
Enzymes are extremely selective, and are often described as adhering to a “lock and key fit” (Figure 1). Their selectivity is primarily enforced by their molecular shape, hydrophilic interactions, and electrostatic forces.
These factors dictate which substrate(s) the enzyme can bind and affect. The activation site is a region on the enzyme where the substrate attaches to form the enzyme/substrate complex. This is also where the reaction occurs. After the enzyme/substrate complex forms and catalysis occurs, the modified substrate is released from the active site, and the enzyme can repeat the process. Enzymes levels are not reduced or altered during the reaction. This means they are efficient and can be used repeatedly.
Figure 1: Demonstration of the “lock and key” fit. The empty pie slice located at the top of Part
A reflects the active site. Part B displays the substrate-enzyme complex. Part C displays the release of the modified substrate, or products.
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Enzymes
Activation Energy
Molecules harbor energy within their chemical bonds. This energy is released when the bonds are broken, thus defining one of the primary reasons why bonds are broken in biological pathways such as metabolism.
However, chemical reactions require an initial amount of energy to break and form bonds (even if the reaction is exothermic). This is often referred to as activation energy as it corresponds to the minimum amount of energy needed to engage (or, activate) a reaction. Enzymes are useful in chemical reactions as they can lower the activation energy required to initiate the reaction. For example, consider trying to roll a large rock on a flat surface. This can be very difficult to do; however, what if you only had to roll the rock down a hill? This would make the process much easier to accomplish. Enzymes are homologous to the downhill slope by decreasing the effort it takes to start a chemical reaction.
Activation energy makes molecules unstable. If sufficient activation energy is available, the bonds existing within the now unstable molecule break to create new, more stable molecules. Activation energy separates the energy levels of the substrates and products and by amounts which are sufficient to initiate the transition state (in which the reactants become unstable) so the reaction can occur. Energy must be added to the substrates to overcome this barrier, which can be recovered when products are formed.
Reaction Kinetics
Enzymes influence the rate at which a chemical reaction occurs. Their activity is affected by three primary factors:
•
Salt concentration
•
Temperature
•
pH
Salt concentrations in the enzyme’s environment, either too high or too low, can influence the behavior of the charged amino acids that make up the protein structure of the enzyme. When this occurs, normal interaction of the charged groups is blocked and the enzyme is no longer functional.
Temperature increases the rate of chemical reactions in two different ways: increasing or decreasing the reaction rate constant and thermal denaturation.
The reaction rate constant can be increased or decreased, and typically adheres to a linear relationship to the direction in which the temperature is adjusted. However, if the temperature rises above an optimum value, enzymes or (their substrates) may become damaged, lose functionality or denature entirely. The pH can influence the efficiency or potential of an enzyme to perform. In fact, the majority of enzymes can only function within a narrow pH range of
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Did You Know...
Enzymes, including proteases and amylases, started being used in laundry detergents in
1913. At the time, this integration was not economically practical, and the enzymes stained the machines used to process the detergent. However, almost 50% of liquid detergents and 25% of powdered detergents now incorporate enzymes to help break down stains which can’t be removed with chemical surfactants alone.
Enzymes approximately 5 - 9. Specifically, pH affects reaction kinematics through the following techniques:
1. Altering the binding site on the enzyme.
2. Changing the protein structure of the enzyme itself
3. Revising the ionization status of the enzyme or substrate.
How do you think the functional pH of enzymes in the stomach compares to the optimal pH for enzymes in blood?
The concentrations of both the enzyme and substrate can also influence the reaction rate (Figure 2). The maximum velocity of the reaction is achieved at a saturating concentration of substrate, at which every enzyme molecule is utilized in the reaction.
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Did You Know...
At low substrate concentrations, an enzyme can readily convert a substrate into products. However, as more substrate is added, an enzyme will reach a saturation point at which the reaction rate will plateau.
This point occurs when there are more substrate molecules than the enzyme can convert in a given period of time. This interaction falls into the category of
“saturation kinetics”.
Figure 2: Reaction rate increases with higher substrate concentrations until it meets maximum velocity.
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Enzymes
Cofactors: Activators and Inhibitors
Substances other than the substrate, called enzyme co-factors, can bind to the enzyme and influence its behavior. Unlike enzymes, coenzymes bind to the enzyme and are structurally modified during the chemical reaction. There are two primary category of cofactors: activators and inhibitors. Activators are chemicals that bind to the active site of the enzyme and help it to bind to the substrate. They are sometimes called cofactors or organic coenzymes.
Inhibitors are chemicals that interfere with the binding of the substrate to the enzyme. There are two types;
1. Competitive: Cannot be replaced by the substrate.
2. Non-competitive: Can not removed by the substrate.
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Did You Know...
Many drugs and poisons are enzyme inhibitors. For example, aspirin inhibits an enzyme that leads to inflammation. Normal cellular processes produce toxic substances (waste) such as hydrogen peroxide and free radicals that will kill the cell if they are not eliminated. Fortunately, yeast and other organisms (including humans) have an enzyme called catalase that breaks down hydrogen peroxide into oxygen and water, which are both harmless to cells. Thus, enzymes are responsible for both constructive
(anabolism) and destructive (catabolism) activities that work synergistically to pattern the overall metabolism of an organism.
Pre-Lab Questions
1. How could you test to see if an enzyme was completely saturated during an experiment?
2. List three conditions that would alter the activity of an enzyme. Be specific with your explanation.
3. Take a look around your house and identify household products that work by means of an enzyme. Name the products, and indicate how you know they work with an enzyme.
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Enzymes
Experiment 1: Enzymes in Food
Amylase is used by humans to facilitate digestion. Specifically, it is an enzyme which breaks down starch molecules into sugar molecules. This is why people sometimes observe a sweet taste after sucking on a starch-containing food for an extended period of time. Amylase is found naturally in human saliva and the pancreas. However, it is also present in some of the common plant foods consumed by humans.
This experiment tests for the presence of amylase in food by using Iodine-Potassium Iodide, IKI. IKI is a color indicator used to detect starch. This indicator turns dark purple or black in color when in the presence of starch. Therefore, if the IKI solution turns to a dark purple or black color during the experiment, one can determine that amylase is not present (because presence of amylase would break down the starch molecules, and the IKI would not change color).
Materials
(1) 2 oz. Bottle (Empty)
(1) 100 mL Graduated Cylinder
*2 Food Products (e.g., ginger root, apple, potato, etc.) 30 mL Iodine-Potassium Iodide, IKI
*Kitchen Knife
Permanent Marker
*Paper Towel
Ruler
*Saliva Sample
2 Spray Lids
*Tap Water
30 mL Liquid Starch, C6H10O5
*Cutting Board
*You Must Provide
Procedure:
1. Remove the cap from the starch solution. Attach the spray lid to the starch solution.
2. Rinse out the empty two ounce bottle with tap water. Use the 100 mL graduated cylinder to measure and pour 30 mL of IKI into the empty two ounce bottle. Attach the remaining spray lid to the bottle.
3. Set up a positive control for this experiment by spraying a paper towel with the starch solution. Allow the starch to dry for approximately one hour (this time interval may vary by local humidity).
4. In the mean time, set up a negative control for this experiment. Use your knowledge of the scientific method and experimental controls to establish this component (hint: what should happen when IKI solution contacts something that does not contain starch?) Identify your negative control in Table 1.
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Enzymes
Note: Be sure to space the positive and negative controls apart from each other to prevent cross contamination.
5. When the starch solution has dried, test your positive and negative controls by spraying them with IKI solution. This step establishes a baseline color scale for you to evaluate the starch concentration of the food products you will test in Steps 7 - 11. Record your results in Table 1.
6. Select two food items from your kitchen cabinet or refrigerator.
7. Obtain a kitchen knife and a cutting board. Carefully cut your selected food items to create a fresh surface.
8. Gently rub the fresh/exposed area of the food items on the dry, starch-sprayed paper towel back and forth 10 - 15 times. Label where each specimen was rubbed on the paper towel with a permanent marker (Figure 3).
9. Wash your hands with soap and water.
10. Take your finger and place it on your tongue to transfer some saliva to your finger. Then, rub your moistened finger saliva into the paper towel. Repeat this step until you are able to adequately moisten the paper towel.
Figure 3: Sample set-up.
Note: You should always wash your hands before touching your tongue! Alternatively, if you do not wish to put your hands in your mouth, you may also provide a saliva sample by spitting in a separate bowl and rubbing the paper towel in the saliva. Be sure not to spit on the paper towel directly as you may unintentionally cross-contaminate your samples.
11. Wait five minutes.
12. Hold the IKI spray bottle 25 - 30 cm away from the paper towel, and mist with the IKI solution.
13. The reaction will be complete after approximately 60 seconds. Observe where color develops, and consider what these results indicate. Record your results in Table 1.
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Enzymes
Table 1: Substance vs. Starch Presence
Substance
Resulting Color
Presence of Starch?
Positive Control: Starch
Negative Control: Student Must Select
Food Product:
Food Product:
Saliva:
Post-Lab Questions
1. What were your controls for this experiment? What did they demonstrate? Why was saliva included in this experiment?
2. What is the function of amylase? What does amylase do to starch?
3. Which of the foods that you tested contained amylase? Which did not? What experimental evidence supports your claim?
4. Saliva does not contain amylase until babies are two months old. How could this affect an infant’s digestive requirements?
5. There is another digestive enzyme (other than salivary amylase) that is secreted by the salivary glands.
Research to determine what this enzyme is called. What substrate does it act on? Where in the body does it become activated, and why?
6. Digestive enzymes in the gut include proteases, which digest proteins. Why don’t these enzymes digest the stomach and small intestine, which are partially composed of protein?
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Enzymes
Experiment 2: Effect of Temperature on Enzyme Activity
Yeast cells contain catalase, an enzyme which helps convert hydrogen peroxide to water and oxygen. This enzyme is very significant as hydrogen peroxide can be toxic to cells if allowed to accumulate. The effect of catalase can be seen when yeast is combined with hydrogen peroxide (Catalase: 2
2 H2O + O2).
H2O2
In this lab you will examine the effects of temperature on enzyme (catalase) activity based on the amount of oxygen produced. Note, be sure to remain observant for effervescence when analyzing your results.
Figure 4: Catalase catalyzes the decomposition of hydrogen peroxide to water and oxygen.
Materials
3 Balloons
3 Test Tubes (Glass)
(2) 250 mL Beakers
Test Tube Rack
30 mL 3% Hydrogen Peroxide, H2O2
Thermometer
Measuring Spoon
Yeast Packet
Permanent Marker
Ruler
*Hot Water Bath
*Stopwatch
20 cm String
*You Must Provide
Procedure
1. Use a permanent marker to label test tubes 1, 2, and 3. Place them in the test tube rack.
2. Fill each tube with 10 mL hydrogen peroxide. Then, keep one of the test tubes in the test tube rack, but transfer the two additional test tubes to two separate 250 mL beakers.
3. Find one of the balloons, and the piece of string. Wrap the string around the uninflated balloon and measure the length of the string with the ruler. Record the measurement in Table 2.
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Enzymes
4. Create a hot water bath by performing the following steps:
a. Determine if you will use a stovetop or microwave to heat the water. Use the 100 mL graduated cylinder to measure and pour approximately 200 mL of water into a small pot or microwave safe bowl (you will have to measure this volume in two separate allocations).
b. If using a stovetop, obtain a small pot and proceed to Step 4c. If using a microwave, obtain a microwave-safe bowl and proceed to Step 4e.
c. If using a stove, place a small pot on the stove and turn the stove on to a medium heat setting.
d. Carefully monitor the water in the pot until it comes to a soft boil (approximately 100 °C). Use the thermometer provided in your lab kit to verify the water temperature. Turn the stove off when the water begins to boil. Immediately proceed to Step 5.
CAUTION: Be sure to turn the stove off after creating the hot water bath. Monitor the heating water at all times, and never handle a hot pan without appropriate pot holders.
e. If using a microwave, place the microwave-safe bowl in the microwave and heat the water in 30 second increments until the temperature of the water is approximately 100 °C. Use the thermometer provided in your lab kit to verify the water temperature. Wait approximately one minute before proceeding to Step 5.
5. Place Tube 1 in the refrigerator. Leave Tube 2 at room temperature, and place Tube 3 in the hot water bath. Important Note: The water should be at approximately 85 °C when you place Tube 3 in it. Verify the temperature with the thermometer to ensure the water is not too hot! Temperatures which exceed approximately 85 °C may denature the hydrogen peroxide.
6. Record the temperatures of each condition in Table 2. Be sure to provide the thermometer with sufficient time in between each environment to avoid obscuring the temperature readings.
7. Let the tubes sit for 15 minutes.
8. During the 15 minutes prepare the balloons with yeast by adding 1/4 tsp. of yeast each balloon. Make sure all the yeast gets settled to the bulb of the balloon and not caught in the neck. Be sure not spill yeast while handling the balloons.
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Enzymes
9. Carefully stretch the neck of the balloon to help ensure it does not rip when stretched over the opening of the test tube.
10. Attach the neck of a balloon you prepared in Step 8 to the top of Tube 2 (the room temperature test tube) making sure to not let the yeast spill into the test tube yet. Once the balloon is securely attached to the test tube lift the balloon and allow the yeast to enter the test tube. Tap the bulb of the balloon to ensure all the yeast falls into the tube.
11. As quickly and carefully as possible remove the Tube 1 (cold) from the refrigerator and repeat steps 9 10 with Tube 1 using a balloon you prepared in Step 8.
12. As quickly and carefully as possible remove Tube 3 (hot) from the hot water bath and repeat steps 9 - 10 with Tube 3 using a balloon you prepared in Step 8.
13. Swirl each tube to mix, and wait 30 seconds.
14. Wrap the string around the center of each balloon to measure the circumference. Measure the length of string with a ruler. Record your measurements in Table 2.
Table 2: Balloon Circumference vs. Temperature
Tube
Temperature (°C)
Balloon Circumference
(Uninflated; cm)
Balloon Circumference
(Final; cm)
1 - (Cold)
2 - (RT)
3 - (Hot)
Post-Lab Questions
1. What reaction is being catalyzed in this experiment?
2. What is the enzyme in this experiment? What is the substrate?
3. What is the independent variable in this experiment? What is the dependent variable?
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Enzymes
4. How does the temperature affect enzyme function? Use evidence from your data to support your answer.
5. Draw a graph of balloon diameter vs. temperature. What is the correlation?
6. Is there a negative control in this experiment? If yes, identify the control. If no, suggest how you could revise the experiment to include a negative control.
7. In general, how would an increase in substrate alter enzyme activity? Draw a graph to illustrate this relationship.
8. Design an experiment to determine the optimal temperature for enzyme function, complete with controls.
Where would you find the enzymes for this experiment? What substrate would you use?
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Lab 9
Cellular Respiration
Cellular Respiration
Learning Objectives
•
Compare and contrast aerobic respiration to anaerobic respiration including the role of ADP and
ATP
•
Relate glucose to pyruvate during the process of glycolysis
•
Explain the role of the citric acid cycle and oxidative phosphorylation in the process of aerobic respiration
•
Explain the role of fermentation in the process of anaerobic respiration
Introduction
Cellular respiration refers to the process of converting chemical energy existing within organic molecules, such as sugars, into a form which can be immediately used by organisms. This process is required for both aerobic (organisms which have oxygen available) and anaerobic (organisms which do not have oxygen available) organisms, but it is executed differently based on the compounds and structures local to the host. However, both processes share the common goal of harvesting biological energy stored in fuel molecules (such as glucose) and convert this energy into ATP. The reaction is written as follows:
C6H12O6
(glucose)
+
6 O2
6 CO2
+
6 H2O +
(oxygen)
(carbon dioxide)
energy
(water)
ADP and ATP
Adenosine triphosphate, or ATP, is often referred to as the energy currency of the cell. It is used in the majority of cellular functions including active transport across a membrane, cell structure maintenance, DNA and RNA synthesis, protein synthesis, etc. However, even though this molecule is extremely important, sometimes more ATP is required than is available.
When ATP levels become too low, a special protein signals the cell to begin respiration.
Respiration occurs in anaerobic and aerobic organisms, therefore, there is more than one pathway used to produce ATP.
However, a high level analysis can conclude that in both types of organisms, insufficient concentrations of ATP can trigger a series of metabolic reactions which ultimately replenish the local
ATP supply. These reactions are primarily catabolic; in other
Figure 1: ATP (above) consists of the nucleotide adenine, a 5-sided sugar (ribose), and three phosphates. Each is bound to three oxygen molecules. Cleavage of one phosphate molecule from ATP yields the amount of energy (calories) found in one peanut.
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Cellular Respiration words, they break down big molecules into smaller pieces. Free energy is released when the molecules are broken down, which can be sequentially used in each proceeding step. Through a series of oxidation reactions, the cell converts carbohydrates into carbon dioxide and water. It also transforms adenosine diphosphate (ADP) into ATP by adding a free phosphate molecule to ADP. This set of reactions provides a constant source of energy for the cell, as long as all the critical components for the reactions are available.
Glycolysis
The first stage of respiration is called glycolysis. Glycolysis occurs with or without oxygen and takes place in the cytoplasm. During glycolysis, glucose is broken down to provide energy for the cell. The glycolytic pathway is highly conserved amongst organisms, and is found in all living organisms.
Glucose contains high energy bonds which release electrons (energy) when broken. Through this process, the six-carbon glucose molecule is broken down into two, three-carbon molecules called pyruvic acid
(pyruvate). Though the bonds holding pyruvate together contain a great deal of potential energy, this step yields little energy. Only two ATP molecules and two nictotinamide adenine dinucleotide (NADH) molecules are produced during glycolysis.
Aerobic Respiration vs. Fermentation
The next stage of respiration depends on if the organism is aerobic or anaerobic; or, in the case of multicellular organisms such as plants, if oxygen is available to the cell. In an aerobic environment, the citric acid cycle begins. Conversely, if an anaerobic environment exists, fermentation or anaerobic respiration takes place.
Plants primarily engage in aerobic respiration. However, in rare cases such as extremely wet soil which results in waterlogged roots, anaerobic respiration may also occur in plants. The steps involved in aerobic respiration, anaerobic respiration, and fermentation vary, but all involve the manipulation of electrons and serve to perpetuate the availability of energy for the cell.
In aerobic respiration, pyruvate molecules are shuttled from the cytoplasm into the mitochondria to prepare for the citric acid cycle. An enzyme complex called pyruvate dehydrogenase complex produces acetylcoenzyme A (acetyl-CoA) by initially breaking down pyruvate to a two-carbon acetyl group and carbon dioxide. The acetyl group is then transferred to coenzyme A. Coenzyme A then acts as a carrier for the twocarbon acetyl group and transfers the acetyl group to the citric acid cycle.
The Citric Acid Cycle
The citric acid cycle, also known as the Krebs cycle or tricarboxylic acid cycle (TCA cycle), is an eight step process in which the acetyl group is oxidized to carbon dioxide. The citric acid cycle engages both anabolic and catabolic reactions to construct and break down large molecules. During this process, two important electron shuttle molecules [flavin adenine dinucleotide (FAD) and nicotinamide adenine dinucleotide (NAD+)] are reduced forming FADH2 and NADH. These molecules shuttle electrons to the next step of aerobic respiration, the electron transport chain.
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Cellular Respiration
The electron transport chain, ETC, performs the final series of biochemical reactions. Ultimately, the
ETC functions to regenerate oxidized molecules (coenzymes) from their reduced state so that other glucose molecules can be converted to energy through future rounds of respiration. To accomplish this objective, electrons (and hydrogen ions) from FADH2 and NADH are transferred to enzyme complexes embedded in the mitochondria’s inner membrane. These complexes act as electron acceptors, passing the electrons from one complex to the next. Since oxygen has a very high affinity for electrons, aerobic respiration is the most efficient means of producing ATP (up to 34 molecules are generated per round).
Figure 2: The electron transport chain.
Fermentation
Eukaryotic and prokaryotic organisms use fermentation in anaerobic conditions. In these cases, the citric acid cycle and the ETC are not used. Instead, pyruvate is the final substrate. These molecules are metabolized in the cytoplasm to produce alcohol or lactic acid, depending on the organism. The main function of fermentation is the reduction of NADH to NAD+. This contributes to the propagation of the glycolytic pathway, which allows ATP production to continue. Recall that glycolysis nets two ATP molecules per glucose molecule. Fermentation allows for the generation of much less ATP per glucose molecule than aerobic respiration because the citric acid cycle and the ETC are not engaged.
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Did You Know...
Yeast has been used to make leavened bread for centuries. When yeast undergoes fermentation, carbon dioxide is trapped between gluten and causes the bread to rise. Ethanol, another byproduct of yeast fermentation, generates the alcohol content in beer, and the carbon dioxide provides effervescence.
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Cellular Respiration
Figure 3: Anaerobic vs. aerobic respiration charts. The green area represents the cytoplasm of the cell where glycolysis and anaerobic respiration occur. The blue area represents the inside of the mitochondria where aerobic respiration occurs.
Industrial Applications
Fermentation has many industrial applications, many of which are evident within the food industry. For example, the sour or tart taste of yogurt and sauerkraut is due to bacteria in the Lactobacillus genus. Lactobacillus perform lactate fermentation in which pyruvic acid (from glycolysis) is reduced directly to the final end product, lactic acid. Another example of industrial fermentation focuses on yeast. Yeast, a unicellular eukaryote, has been used to make leavened (rising) bread for centuries. When yeast undergoes fermentation, carbon dioxide is trapped between gluten and causes the bread to rise. Ethanol is another byproduct of fermentation and is responsible for the alcohol content in beer. Carbon dioxide produced through fermentation is responsible for the effervescence in beer.
Respirometers
Scientists may use respirometers to measure the rate of respiration. This device capitalizes on the ideal gas law for its function. The ideal gas law applies to those gases in which all collisions between atoms or molecules are perfectly elastic and no intermolecular forces exist. The internal energy is completely kinetic, and any change in energy will be accompanied by a change in temperature, volume, and pressure. The relationship is defined by the ideal gas law equation:
PV = nRT
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Cellular Respiration
In this equation, P is the absolute pressure, V is the volume, R is the universal gas constant (8.3145 J/mol °
K), and T is the absolute temperature (Kelvin). Respirometers typically consist of a device to absorb carbon dioxide produced through respiration and measure oxygen uptake by displacement of fluid in a tube connected to the sealed chamber. Since the carbon dioxide is absorbed, air will be taken in through the tube
(displacing the fluid) to maintain equal pressure within the chamber.
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Did You Know...
Cells require more energy during physical activity than when the body is at rest. Humans rely on aerobic respiration to provide this energy. However, some physical activities (such as a strenuous workout) may create an anaerobic environment if the oxygen supply depletes before the demand is removed. When this occurs, lactate fermentation will begin to provide additional energy without the use of oxygen. Lactic acid is a 3-carbon molecule. Therefore, most of the energy remains and the only energy produced is the two ATP molecules generated from glycolysis. The buildup of lactic acid can cause muscles to feel sore and stiff. This feeling persists until the lactic acid is removed from the muscles by diffusion into the blood.
Definitions
• Aerobic Respiration: Oxidation of molecules to produce energy in the presence of oxygen.
•
Anaerobic Respiration: Oxidation of molecules to produce energy in the absence of oxygen.
•
ATP: Adenosine triphosphate; the main energy source of the cell.
•
Citric Acid Cycle (The Krebs Cycle): The step following glycolysis in which acetyl groups produced using pyruvate molecules are oxidized yielding CO2, ATP, H2O and reduced electron shuttle molecules.
•
Fermentation: A metabolic pathway used in anaerobic respiration which reduces NADH to NAD+
•
Glycolysis: The initial process of energy metabolism for anaerobic and aerobic organisms which produces two ATP, two NADH, and two pyruvate molecules.
•
Oxidation: The loss of electrons.
•
Oxidation State: The degree of oxidation of an atom.
•
Pyruvate (Pyruvic Acid): A three-carbon molecule generated through glycolysis.
•
Reduction: Gain of electrons.
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Cellular Respiration
Pre-Lab Questions
1. Why is cellular respiration necessary for living organisms?
2. Why is fermentation less effective than respiration?
3. What is the purpose of glycolysis?
4. How many ATP molecules are produced in aerobic respiration, fermentation, and glycolysis?
Experiment 1: Fermentation by Yeast
Yeast cells produce ethanol, C2H6O, and carbon dioxide, CO2, during alcoholic fermentation. In this experiment, you will measure the production of CO2 to determine the rate of anaerobic respiration in the presence of different carbohydrates with a simplified respirometer.
Materials
(4) 250 mL Beakers
Ruler
15 mL 1% Glucose Solution
15 mL 1% Sucrose (Sugar) Solution, C12H22O11
(1) 100 mL Graduated Cylinder
Test Tube Rack
Measuring Spoon
1 Yeast Packet
1 g. Packets of Equal®, Splenda®, and Sugar
*Stopwatch
Permanent Marker
*Warm Water
3 Pipettes
5 Respirometers (two test tubes that fit into each other – 5 plastic and 5 glass; see Figure 4).
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* You Must Provide
Cellular Respiration
Note: Sucrose (a disaccharide) is made up of glucose and fructose. Glucose is a monosaccharide.
Figure 4: Four respirometers. Note how the smaller, plastic test tube is inverted into the larger test tube. You will create five respirometers in this experiment.
Procedure
1. In this experiment, you will mix yeast with sugar, Equal, and Splenda. Before you begin, develop a hypothesis predicting what will happen when the sugar/sweeteners are mixed with yeast. Example, will fermentation occur? Why or why not? Record your hypothesis in the Post-Lab Questions section.
2. Use the permanent marker to label three 250 mL beakers as Equal®, Splenda®, and Sugar.
3. Empty the Equal®, Splenda®, and Sugar packets into the corresponding beakers.
4. Fill the Equal® and Splenda® beakers to the 100 mL mark with tap water.
5. Fill the Sugar beaker to the 200 mL mark.
6. Mix each beaker thoroughly by pipetting the solution up and down. Each beaker now contains a 1% solution. Set these aside for later use.
7. Completely fill the smallest tube with tap water and invert the larger tube over it. Push the small tube up
(into the larger tube) until the top connects with the bottom of the inverted tube. Invert the respirometer so that the larger tube is upright (there should be a small bubble at the top of the internal tube).
Note: Repeat Step 6 several times as practice. Strive for the smallest bubble possible. When you feel comfortable with this technique, empty the test tube(s) and proceed to Step 7.
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Cellular Respiration
8. Use the permanent marker to label the fourth 250 mL beaker as Yeast.
9. Fill this beaker with 175 mL of warm tap water. The exact temperature does not matter, but the water should be warm to the touch.
10. Open the yeast package, and use the measuring spoon to measure and pour 1 tsp. yeast into the beaker. Pipette the solution up and down until all of the yeast has dissolved into solution.
Note: Make sure the yeast solution remains homogenous before each test tube is filled in the proceeding steps. The yeast concentration is fairly saturated, and yeast may precipitate out of solution if the beaker rests for an extended period of time.
11. Use the permanent marker to label the big and small test tubes as 1, 2, 3, 4, and 5.
12. Use the 100 mL graduated cylinder to measure and pour 15 mL of the following solutions into the corresponding small test tubes:
Tube 1: 1% Glucose Solution
Tube 2: 1% Sucrose Solution
Tube 3: 1% Equal® Solution
Tube 4: 1% Splenda® Solution
Tube 5: 1% Sugar Solution
Note: Rinse the graduated cylinder between each measurement.
13. Fill the remaining volume in each small tube to the top with the yeast solution.
14. Slide the corresponding larger tube over the small tube and invert it as practiced in Step 6. This will mix the yeast and sugar/sweetener solutions.
15. Place the respirometers in the test tube rack, and use a ruler measure the initial air space in the rounded bottom of the internal tube. Record these values in the Table 1.
16. Allow the test tubes to sit in a warm place (approximately 30 °C) for two hours. Placement suggestions include: a sunny windowsill, atop (not in!) a warm oven heated to approximately 85 °C (185 °F on an oven setting), or under a very bright (warm) light.
17. At the end of the respiration period, use your ruler to measure the final gas height (total air space) in the tube. Record this data in Table 1.
Table 1: Yeast Fermentation Data
Tube
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Initial Gas Height (mm)
Final Gas Height (mm)
Net Change
Cellular Respiration
Post-Lab Questions
1. Include your hypothesis from Step 1 here. Be sure to include at least one piece of scientific reasoning in your hypothesis to support your predictions.
2. Did you notice a difference in the rate of respiration between the various sugars? Did the artificial sugar provide a good starting material for fermentation?
3. Was anaerobic fermentation occurring? How do you know (use scientific reasoning)?
4. If you observed respiration, identify the gas that was produced. Suggest two methods you could use for positively identifying this gas.
5. Hypothesize why some of the sugar or sweetener solutions were not metabolized, while others were. Research the chemical formula of Equal® and Splenda® and explain how it would affect yeast respiration.
6. How do the results of this experiment relate to the role yeast plays in baking?
7. What would you expect to see if the yeast cell metabolism slowed down? How could this be done?
8. Indicate sources of error and suggest improvement (for example, what types of controls could be added?).
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Cellular Respiration
Experiment 2: Aerobic Respiration in Beans
We will evaluate respiration in beans by comparing carbon dioxide production between germinated and nongerminated beans. As shown in the balanced equation for cellular respiration, one of the byproducts is CO2
(carbon dioxide):
C6H12O6 + 6 H2O + 6 O2
energy + 6 CO2 +12 H2O
We will use a carbon dioxide indicator ( bromothymol blue) to show oxygen is being consumed and carbon dioxide is being released by the beans. Bromothymol blue is an indicator that turns yellow in acidic conditions, green in neutral conditions, and blue in basic conditions. When carbon dioxide dissolves in water, carbonic acid is formed by the reaction:
H2O + CO2
H2CO3
resulting in the formation of this weak acid. If an indicator such as bromothymol blue is present, what do you think would happen? (Hint - what color would the indicator change to?)
Materials
(6) 250 mL Beakers
1 Pipette
24 mL Bromothymol Blue Solution, C27H28Br2O5S
6 Rubber Bands (Large. Contains latex; please handle with gloves if you have a latex allergy.)
100 Kidney Beans
6 Medicine Cups (small, clear, plastic cups)
15 cm Parafilm®
*Water
*Paper Towels
Permanent Marker
100 Pinto Beans
*You Must Provide
Procedure
1. Label two of the 250 mL beakers as Soaked: Pinto and Soaked: Kidney.
2. Fill each beaker with 200 mL water.
3. Count and transfer 50 pinto beans into the Soaked: Pinto beaker. Then, count and transfer 50 kidney beans into the Soaked: Kidney beaker. Allow the beakers to rest for 24 hours.
4. After 24 hours have passed, carefully strain the water from each beaker.
5. Place two paper towels on a flat work surface. Use the permanent marker to label one paper towel as
Soaked: Pinto and the second as Soaked: Kidney.
6. Pour the soaked beans onto paper towels, keeping them sorted by bean type.
7. Label the remaining beakers as Dry: Pinto, No Pinto, Dry: Kidney, and No Kidney.
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Cellular Respiration
8. Place several layers of moist paper towels at the bottom of all six, 250 mL beakers.
9. Place 50 pre-soaked pinto beans into the Soaked: Pinto beaker, 50 unsoaked pinto beans into the Dry:
Pinto beaker, and no beans into the No Pinto beaker.
10. Place 50 pre-soaked kidney beans into the Soaked: Kidney, 50 unsoaked kidney beans into the Dry: Kidney beaker, and no beans into the No Kidney beaker.
11. Dispense four mL of bromothymol blue solution into each of the six measuring cups. Then, place one cup inside each beaker (Figure 5).
12. Stretch Parafilm® across the top of each beaker. Secure with a rubber band to create an air-tight seal.
Note: If your Parafilm® seal breaks, plastic wrap
(such as Saran® wrap) can be used as a replacement.
13. Place the beakers on a shelf or table, and let sit undisturbed at room temperature.
14. Record the initial color of the bromothymol solutions and observe the jars at 30 minute intervals for three hours. Record the color of the bromothymol blue in
Tables 2 and 3.
Figure 5: The image above shows what the beans,
15. Let the beans and the jar sit overnight. Record your beaker, and measuring cup set-up should look like. final observations in Tables 2 and 3.
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Cellular Respiration
Table 2: Bromothymol Blue Color Change Over Time for Pinto Bean Trial
Time
Pre-Soaked Pinto Beans
Dry Pinto Beans
No Pinto Beans
0 min
30 min
60 min
90 min
120 min
150 min
180 min
24 hours
Table 3: Bromothymol Blue Color Change Over Time for Kidney Bean Trial
Time
0 min
30 min
60 min
90 min
120 min
150 min
180 min
24 hours
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Pre-Soaked Kidney Beans
Dry Kidney Beans
No Kidney Beans
Cellular Respiration
Post-Lab Questions:
1. How did the color of the bromothymol blue solution in each beaker change over time in each condition?
2. What is the mechanism driving the bromothymol blue solution color change?
3. What can be inferred from the color change of the bromothymol blue solution?
4. What evidence do you have to prove that cellular respiration occurred in the beans? Explain your answer.
5. What are the controls in this experiment, and what variables do they eliminate? Why is it important to have a control for this experiment?
6. If this experiment were conducted at 0 °C, what difference would you see in the rate of respiration? Why?
7. Would you expect to find carbon dioxide in your breath? Why?
8. What else could you incorporate into this experiment to verify that the gas is responsible for the color change? Design an experiment that shows the steps required.
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Lab 10
Cell Structure and Function
Cell Structure and Function
Learning Objectives
•
Apply Cell Theory
•
Compare and contrast the structure and function of prokaryotic and eukaryotic cells
•
Identify eukaryotes and prokaryotes based on their cellular structure
Introduction
A cell is the fundamental unit of life. All living organisms originate from a single cell. Some remain as a single cell, while others grow and divide to form a multi-cellular organism. Though most cells are difficult to see with the naked eye, cytologists have successfully identified many of their features using microscopes. The scope of these features ranges from the fundamental characteristics of the outer membranes, to the internal structures nestled within the cytoplasm. This structural information provided the foundation for much of modern biology, and formed the basis of Cell Theory.
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Cell Theory states:
• All living things are made of cell(s).
• A cell is the smallest unit of life.
Did You Know...
Cytologists are scientists who study cells.
The study of the cell is known as cytology.
• All cells are generated from previous cells.
• All cells pass on genetic information.
• Energy metabolism occurs within cells.
• The chemical make-up of all cells is similar (but not identical).
Functional Requirements of Life
The functional requirements of life are similar to the content presented in Cell Theory. In essence, these requirements state that all living organisms (unicellular or multicellular) must perform the following functions:
1. Movement: Cells can change position or shape.
2. Responsiveness: Cells can sense internal or external changes; and, react to these changes.
3. Respiration/Metabolism: Cells produce energy through cellular chemical reactions
4. Reproduction: Cells are able to produce offspring.
5. Excretion: Cells can remove waste produced by metabolic activities.
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Cell Structure and Function
Types of Cells: Prokaryotes and Eukaryotes
Although all organisms are made up of one or more cells, not all cells are identical. Prokaryotes and eukaryotes are organisms composed of two structurally different types of cells. This division offers the most broad distinction between different types of organisms. Prokaryotes are the most ancient and basic organisms, and span the taxonomic classes of bacteria and archaea. They lack a membrane-bound nucleus and membranebound organelles (specialized structures). The term prokaryote comes from the Latin words “pro” (before) and
“karyote” (nucleus).
Eukaryotes are much more complex organisms with two characteristics that set them apart from prokaryotes: a defined nucleus, and membrane-bound organelles. The term “eukaryote” comes from the Latin words
“eu” (true) and “karyote” (nucleus). Fungi, plant, protist, and animal cells are all examples of eukaryotic cells.
Prokaryotic and Eukaryotic Structural Similarities
While the overall structure of prokaryotic and eukaryotic cells differs, there are several structures which are commonly found in all cells. For example, all cells have deoxyribonucleic acid (DNA), cytoplasm, a cell membrane, and ribosomes.
DNA
DNA contains all the genetic information for the cell. It is made up of two long chains of nucleotides. Each nucleotide consists of a base, a five carbon sugar molecule, and at least one phosphate group. Adenine, thymine, guanine, and cytosine are the four different types of nucleotides. These nucleotides connect to form a sugar phosphate backbone, and extend hydrogen bonds out towards complimentary DNA strands to form a double helix structure. The double helix is often compared to a long, twisted ladder. DNA is often found as a DNA-protein complex known as chromatin. Chromatin condenses during cell cycle division to form chromosomes.
G
C
A
T
C
G
T
A
Figure 1: The hydrogen bonds formed
All cells have the DNA described, although eukaryotic DNA is linear, between adenine and thymine and while prokaryotic DNA is circular. Also, because prokaryotes do not cytosine and guanine contribute to have a nucleus, their DNA exists freely within the nucleoid region as the double-helix structure of DNA opposed to the eukaryotic location within the nucleus.
Cytoplasm
Cytoplasm is a fluid-like substance which also exists in all cells. The cytoplasm is made up cytosol, a water based solution containing small molecules, such as proteins and sugars. Because of the abundance of small
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Cell Structure and Function molecules and particles, the cytoplasm is better understood as a thicker, gel-like solution instead of a highly viscous liquid solution. All of the everyday cellular functions are carried out in the cytoplasm, including cell replication and cell growth.
There are also many biomolecules, such as ribosomes, that are suspended within the cytoplasm. Ribosomes are particles made up of RNA and proteins and are roughly 20 nm (typical prokaryotic length) to 30 nm (typical eukaryotic length) in diameter. These particles are the site for the complex process of protein synthesis in all cells.
Cell Membrane
The cytoplasm is surrounded by a cell membrane. The cell membrane provides structural support and strength. It also protects the cell and the cellular content from the external environment. It acts as a selective gateway which regulates the flow of atoms and molecules in and out of the cell. The cell membrane can be described by the fluid mosaic model which states that cell membranes are not static. Instead, they express movement by the molecules that compose it. Cell membranes are often referred to as a phospholipid bilayer, as they are composed of two layers of phosphate-containing lipids with proteins floating between these layers. This forms through the process of “self assembly” by which the hydrophobic tails of phospholipid molecules secure an inward position within the membrane. This prevents it from contacting water on a normal basis. Conversely, the hydrophilic heads of the phospholipids are positioned outward and are in contact with water. The proteins in the structure are responsible for carrying out the majority of the functions specific to the membrane and imparts the selectivity to certain materials that can pass through the membrane.
Figure 2: A phospholipid bilayer (cell membrane) with embedded proteins.
Cell Wall
Many cells within the prokaryotic and eukaryotic families also have cell walls outside the cell membrane that help to protect them and provide strength and support; although, animal cells and protozoa do not have cell walls. Unlike the cell membrane, the cell wall is not selective and does not allow materials to pass through easily. Prokaryotic cells have a thick, rigid cell wall composed of amino acids and sugars (peptidoglycan), but the cell wall composition within eukaryotes varies. For example, fungi cell walls include a polysaccharide called chitin while plants exhibit cell walls with the polysaccharide cellulose.
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Cell Structure and Function
Prokaryotes and eukaryotes must also regulate nutrients and wastes, and require a supply of energy to exist.
Metabolic activities such as photosynthesis and respiration can be carried out by both cell types. However, photosynthetic activity initiates in the chloroplasts in eukaryotes, while it occurs in the thylakoid in prokaryotes. The thylakoid is an extension of the plasma membrane that contains photosynthetic pigments.
Prokaryotic Characteristics
Pili
Both physical and chemical diversity can be viewed between prokaryotic and eukaryotic cells. A structural examination can begin by reviewing major prokaryotic structures, such as the flagella, although prokaryotes tend to have fewer organelles and diverse features than eukaryotes. Flagella are long, cylindrical protrusions on the outside of a cell. They are often, but not always, present, and facilitate mobility through a whiplike, motor rotation. For example, E. coli bacteria rotate their flagellum in a clockwise direction to swim through
Cell Wall their environment.
Ribosomes
Nucleoid
Region
Some prokaryotic cells also have a glycocalyx, which Cytoplasmic is a slime-coating used to protect the cell. The gly- Membrane cocalyx also allows prokaryotes to attach to different surfaces (such as teeth, lungs, or even artificial joints).
This is helpful as these surfaces provide a nutritional resource and often a safe habitat.
Cytoplasm
Many prokaryotic bacteria cells also posses pili. Pili are small, hair-like structures on the outer surface of many
Flagella
types of bacteria that provide the ability to adhere to hosts and other bacteria, as well as the ability to trans- Figure 3: Prokaryotic cell diagram showing some of mit genetic information. Bacterial cells that posses sex the major structures including flagella, pili, freely pili have the ability to transmit genetic information with existing DNA, and ribosomes. other cells. This may result in the transmission of genes that provide antibacterial (antibiotic) resistance.
Eukaryotic Characteristics
Eukaryotic cells are generally more structurally complex than prokaryotic cells and typically contain more components. A network of membrane-bound organelles exists within each eukaryotic cell. Essentially, these
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Cell Structure and Function organelles function as “mini” organs within the cell. Each organelle fulfills a
Did You Know... unique function, but they must all work together to maintain homeostasis within the cell.
Red blood cells are created in
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Nucleus
The nucleus is often referred to as the control center of the cell. It contains nuclear DNA, which codes for the majority of the proteins required for human survival. The nucleus is surrounded by a double membrane system, called the nuclear envelope. This structure contains nuclear pores that facilitate communication between the nucleus and other cellular structures via active and passive transport of molecules and ions. A smaller, dense structure called the nucleolus also exists within the nucleus. The nucleolus is composed of RNA and proteins. This structure functions to create the ribosomal
RNA (rRNA) and assemble necessary ribosomes within the cell. The exact size of this structure is dependent upon the necessary amount of ribosomes.
For example, if great need for ribosomes exists within the cell, the nucleolus is typically larger. Conversely, the nucleolus remains smaller if only a small number of ribosomes is required.
the bone marrow. Once they mature and enter the blood stream their nucleus degrades and they live the rest of their lives without one.
This frees up space to carry oxygen and makes it easier to fit through small capillaries.
The Mitochondrion
While most eukaryotic DNA is housed within the nucleus, mitochondrial DNA is located in a cell organelle called the mitochondrion. Mitochondria (more than one mitochondrion) are separate organelles located in the cytoplasm, and are often considered the powerhouse of the cell because of their functional involvement in energy metabolism. In fact, this small, elongated structure is the site for cellular respiration in which nearly all of the energy of the cell is produced (when oxygen is present). Mitochondrial DNA enFigure 4: Mitochondrion. codes genes which aid in energy production. Mitochondria are housed within a double membrane structure. The outer membrane serves to hold and protect the organelle while the inner membrane, called the cristae, contains multiple folds that allow for a greater surface area.
The increased surface area allows for more efficient energy production. There is always at least one mitochondrion in eukaryotic cells. However, cells which require a great amount of energy, such as muscle cells, may contain thousands of individual mitochondria.
The Cytomembrane System
Within the entire network of the cell lies the cytomembrane system composed of the endoplasmic reticulum, golgi body and many vesicles. This system is responsible for the production of lipids and proteins. The
147
Cell Structure and Function cytomembrane system begins in the endoplasmic reticulum (ER) which, in animal cells, is connected directly to the outer layer of the nuclear envelope. The
ER is divided into two components, the rough ER and the smooth ER. The rough ER is composed of flattened sac-like structures with attached ribosomes and functions in protein synthesis and protein transportation to the golgi body. The term “rough ER” is derived from this rough appearance. The surface appears bumpy or rough due to the ribosomes which are attached to it. The smooth ER is also composed of flattened sac-like structures, and is devoid of ribosomes.
Thus, the smooth ER appears smooth under a microscope. The smooth ER functions to create many of the lipids of the cell and also functions in detoxification of destructive chemicals that may pass throughout the cell. 2
1
3
Figure 5: The cytomembrane system. Note the endoplasmic reticulum (1) sprouting from the outer layer of the nuclear envelope (2), and the flattened, saclike appearance of the golgi apparatus (3) in the lower left.
The next step in the cytomembrane system is the Golgi body, also called the Golgi apparatus. This structure is composed of flattened sac-like structures stacked on top of each other. The proteins and lipids created in the ER are transferred to the Golgi body where they are modified, packed and then shipped out to specified locations both within and outside of the cell.
After the proteins and lipids are modified and packed in the Golgi body, they are then transported in vesicles. One type of vesicle, called the secretory vesicle, is composed of a sac freed or pinched off of the Golgi body. Secretory vesicles transport proteins and lipids to the plasma membrane so they can be released or secreted from the cell.
Another type of vesicle, the lysosome, is also released off the Golgi body. These vesicles contains enzymes that are used in the digestion of cellular components. They can digest proteins, lipids, carbohydrates, cell parts and even an entire cell.
Another membrane bound vesicle is the peroxisome. The peroxisome is responsible for lipid and amino acid break down, as well as the break down of hydrogen peroxide, a highly toxic byproduct of certain cellular reactions. When drinking alcohol, peroxisomes also function to break down almost half of all that was consumed.
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Cell Structure and Function
The Cytoskeleton
The cytoskeleton is a network of interconnected fibers within the cell that provides shape, internal organization, and the ability for a cell to move. Microtubules are a part of the cytoskeleton of a cell. These hollow cylinders are composed of tubulin protein and have a diameter of about 25 nm. Microtubules are key in cellular structure, providing support in the cytoplasm. They are also very important in cellular division as they form the centrioles and spindle fibers necessary for the cells to divide. Microtubules can also function in the movement of the cell by combining to form flagella and cilia. Flagella are long, cylindrical protrusions on the outside of a cell. They provide mobility through a whip-like rotation. Cilia are much smaller, hair-like and typically more abundant than flagella. The cilia beat together to either allow the cell to move or to cause movement of extracellular fluid.
The side kick to the microtubules are the microfilaments, also known as actin filaments. Microfilaments are much smaller and thinner than their microtubule counterpart, but are widely abundant throughout the cell.
They are made of the protein actin and are typically no larger than 8 nm (Remember: 1 nanometer (nm) = 1 x 10-9 meter) in diameter. These cellular components are also responsible for cellular structure within the cytoplasm along with cellular movement. Another support component within the cytoplasm is the intermediate filaments. These filaments are about 10 nm in diameter and provide additional strength to the cytoplasm of the cell. There are many cellular components that work together to facilitate all the responsibilities of a cell.
Each individual component works to perform its specific function so that the cell, as a whole, can perform the major functions required of it.
Figure 6: Parts of the cytoskeleton.
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Cell Structure and Function
Animal Cell (top); Plant Cell (bottom)
150
Figure 7: Animal and plant cells are both eukaryotic.
Cell Structure and Function
Quick Reference: Eukaryotic Organelles
•
Cytoskeleton: The cell “skeleton” found in all eukaryotic cells that provides shape to the cell while also enabling it to move.
•
Endoplasmic Reticulum (ER): A series of membranes extending throughout the cytoplasm that can be peppered with ribosomes (rough ER) or not (smooth ER). The rough ER functions in protein synthesis while the smooth ER functions in lipid synthesis.
•
Golgi Apparatus (also called the Golgi Body): A series of flattened sac-like bodies that processes the cell’s proteins and lipids before they are released to their final destination.
•
Lysosomes: Enzyme-filled vesicles found within the cell that aid in intracellular digestion.
•
Mitochondrion (plural: mitochondria): The “power plant” of the cell. These are membrane bound organelles (inner and outer membrane) with their own circular DNA, and make ATP (energy) for the rest of the cell.
•
Nuclear Envelope: An outer membrane that surrounds the nucleus.
•
Nuclear Pores: Holes in the nuclear envelope that permit communication between the internal nuclear environment and the cytoplasm.
•
Nucleus: Houses the genetic content (DNA) of the cell.
•
Nucleolus (plural: nucleoli): A part of the nucleus that is made of RNA, protein and chromatin; and, manufactures rRNA and ribosomes.
•
Peroxisomes: Contain enzymes that help the cell destroy toxins.
•
Ribosomes: Ribosomes are large molecules found in all living cells. Ribosomes the site of protein synthesis. A strand of mRNA docks onto a ribosome. The correct amino acids are then recruited to the ribosome to create a protein.
•
Vacuole: A fluid-filled organelle that helps isolate its contents from the cytoplasm.
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Cell Structure and Function
Table 1: Structural Variations Between Prokaryotic and Eukaryotic Cells
Structure
Present in Prokaryotic Cell?
Present in Eukaryotic Cell?
Nucleus
No
Yes
Plasma Membrane
Yes
Yes
Cell Wall
Yes
Yes (plant and fungal cells; not animal cells)
Cytoplasm
Yes
Yes
Flagella
Occasionally
Occasionally
Pili
Occasionally
No
Cilia
No
Occasionally
Glycocalyx
Occasionally
Occasionally
Cytoskeleton
No
Yes
Endoplasmic Reticulum
No
Yes
Mitochondria
No
Yes
Golgi Apparatus
No
Yes
Chloroplast
No
Yes, in plants and other photosynthetic organisms
Ribosome
Yes
Yes
Lysosome
No
Yes
Peroxisome
No
Yes
Vacuole and Vesicle
Occasionally
Yes (in most cells)
Pre-Lab Questions
1. Identify three major similarities and differences between prokaryotic and eukaryotic cells.
2. Where is the DNA housed in a prokaryotic cell? Where is it housed in a eukaryotic cell?
3. Identify three structures which provide support and protection in a eukaryotic cell.
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Cell Structure and Function
Experiment 1: Identifying Cell Structures
The structure of a cell dictates the majority of its function. You will view a selection of slides that exhibit unique structures that contribute to tissues function.
Materials
Onion Root (allium) Digital Slide Images
Procedure
1. Examine the onion root tip digital slide images on the following pages. Then, respond to the Post-Lab
Questions.
Onion Root Tip: 100X
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Cell Structure and Function
Cytoplasm
Chromosomes
Chromosomes
Cell wall
Nucleus
Onion Root Tip: 1000X
Cytoplasm
Cell wall
Nucleus
Chromosomes
154
Onion Root Tip: 1000X
Cell Structure and Function
Onion Root Tip: 100X. Each dark circle indicates a different nucleus.
Cell wall
Cytoplasm
Chromosomes
Onion Root Tip: 1000X
Nucleus
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Cell Structure and Function
Post-Lab Questions
1. Label each of the arrows in the following slide image:
A
C
D
Onion Root Tip: 1000X
B
2. What is the difference between the rough and smooth endoplasmic reticulum?
3. Would an animal cell be able to survive without mitochondria? Why or why not?
4. What could you determine about a specimen if you observed a slide image showing the specimen with a cell wall, but no nucleus or mitochondria?
5. Hypothesize why parts of a plant, such as the leaves, are green, but other parts, such as the roots, are not. Use scientific reasoning to support your hypothesis.
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Cell Structure and Function
Experiment 2: Create a Cell
In this experiment, you will create an animal and a plant cell using household items to observe the difference between prokaryotes and eukaryotes.
Materials
30 cm x 30 cm Aluminum Foil (Cell Wall)
2 Resealable Bags (Cell Membrane)
20 Beads, any color (Rough Component of Endo- *Bowl plasmic Reticulum and free ribosomes)
*Household items to represent the Golgi bodies
2 Eggs (plastic) (Nucleus)
and chloroplasts
4 Gelatin Packets, unflavored
*Warm Water
12 Kidney Beans (Mitochondria)
Permanent Marker
*You Must Provide
4 Pipe Cleaners (Endoplasmic Reticulum)
Procedure
1. Place four packets of unflavored gelatin in a bowl. Add 4 cups of hot water to the bowl. Do not refrigerate the mixture yet!
Note: You do not need to heat the water in a microwave. Simply run tap water until it feels warm to the touch.
2. Label each resealable bag as either “Plant Cell” or “Animal Cell”. These will serve as the cell membrane.
3. Construct rough endoplasmic reticulum by stringing 10 beads onto two of the pipe cleaners (5 on each). The remaining, unbeaded pipe cleaners represent smooth endoplasmic reticulum.
4. Construct a cell wall using the aluminum foil. This should be large enough to fit the resealable bag when filled with half of the gelatin and some of the cell structures.
Hint: It is helpful to make this square-shaped.
5. Using your knowledge of the Golgi apparatus and chloroplasts, think of two household items which can represent these structures. Find and collect these items for use in this experiment.
Hint: Colored paper may bleed when placed in gelatin.
6. Open the resealable bag labeled “Plant Cell” and pour half of the liquid gelatin into it.
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Cell Structure and Function
7. Add the items which represent plant cell structures (you must determine which items!) into the gelatin and tightly close the bag. If there is an “organelle” present in both plant and animal cells make sure to leave enough to be included in the animal cell.
8. Place the bag in the aluminum foil cell wall.
9. Open the resealable bag labeled “Animal Cell” and pour the remainder of the gelatin into it.
10. Add the items which represent animal cell structures (you must determine which items!) into the gelatin and tightly close the bag.
11. Place both “cells” into the refrigerator for 24 hours.
12. Return after 24 hours and observe the “cells” you have made. Notice the difference between the animal cell and the plant cell.
Post-Lab Questions
1. What cell structures did you place in the plant cell that you did not place in the animal cell?
2. Is there any difference in the structure of the two cells?
3. What structures do organisms that lack cell walls have for support?
4. How are organelles in a cell like organs in a human body?
5. How does the structure of a cell suggest its function? List three examples.
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Lab 11
Mitosis
Mitosis
Learning Objectives
•
Relate DNA to genes, alleles and chromosomes
•
Identify chromosomal structure during the various stages of the cell cycle including chromatin, sister chromatids, homologous chromosomes, and centromeres
•
Compare and contrast animal and plant mitosis
•
Apply the concept of ploidy to the human species
•
Explain the role of parent and daughter cells in cell cycle division
Introduction
All of a cell’s hereditary information is encoded by the genome. The genome, all of an organism’s genes, is vital for life as it directs the formation of all the proteins the cell needs to survive. It also allows individual cells to work together and form complex, multicellular organisms. However, both unicellular and multicellular organisms must be able to grow and divide for life to continue. Cell cycle division provides a mechanism by which a cell can pass along its DNA from one generation to the next, and ensure species propagation. Mitosis is the term for eukaryotic cell division, while binary fission describes prokaryotic division.
DNA and Genes
In eukaryotes, the genome is encoded by chromatin, which is composed of deoxyribonucleic acid (DNA), ribonucleic acid
(RNA), and proteins. DNA is a double-stranded molecule, constructed from repeating units of a phosphate molecule, a sugar (deoxyribose), and one of four possible nucleic acids adenine, thymine, cytosine, or guanine. Units which incorporates these three structures are called nucleotides. Hydrogen bonds between complementary nucleotides (adenine and thymine [A - T] and cytosine to guanine [C - G]) hold DNA strands together. Sections of DNA that code for traits such as eye color are called genes (Figure 1).
Ploidy
Figure 1: Chromosomes are long strands of
DNA coiled up. Pieces of DNA that code for traits are called genes.
The number of genes required for an organism to survive varies by species. In fact, different species may have a different number of chromosomes, as well as a different number of copies of each chromosome. Somatic (any body cells, but sex cells) human cells have two copies of 23 chromosomes, resulting in 46 chromosomes per cell. These cells are considered diploid, because they have two copies.
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Mitosis
Almost all somatic, mammalian cells have two complete sets of chromosomes. One set originates from the female parent and one set originates from the male parent, resulting in pairs of homologous chromosomes
(Figure 2).
Cell Cycle Division
Cell cycle division occurs when a parent cell copies its entire genome and splits into two identical daughter cells. Do not confuse the word
“parent” with mother or father. It is just a term to differentiate the cell that is splitting from the two new cell created. Each new daughter cell (not to
Figure 2: Sister chromatids (a) and be associate with the female sex) contains two complete sets of chromohomologous chromosomes (b) of somes. The cellular content is identical to the parent cell as long as no a diploid organism in which 2n=4. mutations or errors occur during cell cycle division.
Cell cycle division is divided into three phases: interphase, mitosis, and cytokinesis (Figure 3). These phases are further broken down into individual stages. These stages are identified as:
Interphase; Stage:
•
Gap 1 (G1): The first stage of interphase; or, the first growth phase.
•
S: DNA synthesis. The second stage of interphase.
•
Gap 2 (G2): The third stage of interphase; or, the second growth stage.
Mitosis; Stage:
•
Prophase
•
Metaphase
•
Anaphase
•
Telophase
Cytokinesis:
•
Figure 3: Mitosis is only one stage of the cell cycle.
There are no further subdivided stages within cytokinesis.
A cell normally completes the cycle in 18 - 24 hours. Mitosis only occupies one to two hours of that period.
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Mitosis
Each stage is regulated by specialized proteins, such as cyclins and cyclin-dependent kinases (CDKs) that coordinate the division and cell growth. Certain types of cancer are associated with the failure of these proteins.
During interphase, the cell prepares for mitosis and cytokinesis by growing larger in size, duplicating its internal organelles (G1), replicating its DNA (S) and preparing for cell division (G2).
You can easily remember the activity which occurs during the S phase because it is when new DNA is synthesized (synthesized starts with s). When the full set of DNA is replicated, two identical copies of each chromosome exist in the parent cell. These identical copies pair up and are referred to as sister chromatids. Sister chromatids exist as loose strands of genetic content known as chromatin during interphase.
These structures don’t condense into chromosomes until prophase of mitosis. To form a chromosome, two sister chromatids are joined together as a four-arm structure by a centromere (see prophase stage below).
Chromosomes also incorporate a variety of proteins, such as histones, which tightly package the DNA. Interestingly, if the chromatin from a single human cell was uncoiled and stretched out, it would be over two meters long. However, it is important that it remain in a loose configuration to permit transcription of DNA.
After the DNA is fully replicated, the cell engages in “proofreading” and additional protein production (G2). In this process, proteins involved in DNA replication (such as DNA polymerase) review the replicated content and correct any errors which may have been made during replication. G1 and G2 are also responsible for initiating a variety of checkpoints which verify that the size and environmental growth factors are sufficient prior to entering mitotic division.
Mitosis follows interphase. It is during mitosis that sister chromatids segregate into two daughter cells. The four stages of mitosis are described below:
Mitosis (Figure 2):
•
Prophase: Loosely formed chromatin condenses into tightly packed chromosomes. Centromeres attach themselves to coiled (condensed) sister chromatids to hold their structure together.
The nuclear envelope of the parent cell breaks down. Pairs of centrioles, which serve as cellular anchors during division, appear at the mitotic spindle, located at the cellular poles at separate sides of the cell.
•
Metaphase: Sister chromatids migrate towards the metaphase plate. Microtubules (long strands) grow from the centrioles and link together while attaching to each pair of sister chroma-
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Mitosis tids. The orientation of each pair of sister chromatids is independent from all other pairs. This means they can “flip flop” as they line up, effectively shuffling their genetic information into new combinations. •
Anaphase: The microtubules attached to the centromeres shorten This causes the sister chromatids to separate and move toward opposite poles of the parent cell. At this point, each sister chromatid is now an individual chromosome. Microtubules attached to other microtubules from the opposite pole lengthen, effectively elongating the cell.
•
Telophase: One set of chromosomes arrives at each pole. A new nuclear envelope begins to form, chromosomes are uncoiled back into chromatin, and a new nucleus is formed at each end of the cell.
Cell cycle division concludes with cytokinesis. During animal cell cytokinesis, the plasma membrane of the cell folds in and encloses around each nucleus, creating two, diploid daughter cells.
Mitosis is virtually identical in plant and animal cells, however, there is one small difference which occurs during telophase. Plants, due to the presence of a cell wall, cannot “pinch” the cytoplasm into two daughter cells. Instead, a new cell wall must be developed which will then separate the two cells, allowing them both to be fully covered with a cell wall.
Prophase
Metaphase
Anaphase
Telophase
Figure 4: The stages of mitosis.
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Cytokinesis
Mitosis
Definitions
Alleles – Alternate forms of the same gene.
Centriole – An organelle that forms the microtubules which separate chromosomes during cell division. Centromere – The region of the chromosome where chromatids are held together. This forms the kinetochore. This is also where microtubules attach during cell division.
Chromatids – One of two identical chromosomes (often called “sister chromatids”).
Chromosome – A threadlike structure consisting of chromatin that contains a long, continuous strand of DNA and its associated proteins.
Cytokinesis – The final step of the cell cycle where the cell splits to produce two daughter cells.
Diploid – Cells that have two copies (2n) of each chromosome.
Gene – A sequence of DNA that codes for a specific detectable product such as a protein or
RNA.
Genetic Variation – The diversity in the DNA found in otherwise similar organisms.
Homologous Chromosome – Chromosomes that contain the same genes, but potentially different alleles.
Locus – The exact position of a gene on a chromosome.
Microtubules – Long polymer strands of protein that separate chromosomes during cell division.
Ploidy – The number (n) of copies of each chromosome contained in a cell (usually 1n or 2n in animals). Tetrad – Four homologous chromosomes, two of each pair of sister chromatids, that line up during prophase I in meiosis.
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Mitosis
Pre-Lab Questions
1. What are chromosomes made of?
2. Research the differences that exist between mitosis and binary fission. Identify at least one difference, and explain why it is significant.
3. Cancer is a disease related to uncontrolled cell division. Investigate two known causes for these rapidly dividing cells and use this knowledge to invent a drug that would inhibit the growth of cancer cells.
Experiment 1: Observation of Mitosis in a Plant Cell
In this experiment, we will look at the different stage of mitosis in an onion cell. Remember that mitosis only occupies one to two hours while interphase can take anywhere from 18 - 24 hours. Using this information and the data from your experiment, you can estimate the percentage of cells in each stage of the cell cycle.
Materials
Onion (allium) Root Tip Digital Slide Images
Procedure
Part 1: Calculating Time Spent in Each Cell Cycle Phase
1. The length of the cell cycle in the onion root tip is about 24 hours. Predict how many hours of the 24 hour cell cycle you think each step takes. Record your predictions, along with supporting evidence, in
Table 1.
2. Examine the onion root tip slide images on the following pages. There are four images, each displaying a different field of view. Pick one of the images, and count the number of cells in each stage. Then count the total number of cells in the image. Record the image you selected and your counts in Table 2.
3. Calculate the time spent by a cell in each stage based on the 24 hour cycle:
Hours of Stage = 24 x Number of Cells in Stage
Total Number of Cells Counted
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Mitosis
Part 2: Identifying Stages of the Cell Cycle
1. Observe the images of the root cap tip.
2. Locate a good example of a cell in each of the following stages: interphase, prophase, metaphase, anaphase, and telophase.
3. Draw the dividing cell in the appropriate area for each stage of the cell cycle, exactly as it appears. Include your drawings in Table 3.
Nucleus
Cytoplasm
Chromosomes
Cell wall
Onion Root Tip: 100X
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Mitosis
Onion Root Tip: 100X
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Onion Root Tip: 100X
Mitosis
Onion Root Tip: 100X
Table 1: Mitosis Predictions
Predictions:
Supporting Evidence:
Table 2: Mitosis Data
Number of Cells in Each Stage
Total Number of Cells
Calculated % of Time Spent in Each Stage
Interphase:
Interphase:
Prophase:
Prophase:
Metaphase:
Metaphase:
Anaphase:
Anaphase:
Telophase:
Telophase:
Cytokinesis:
Cytokinesis:
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Mitosis
Table 3: Stage Drawings
Cell Stage:
Interphase:
Prophase:
Metaphase:
Anaphase:
Telophase:
Cytokinesis:
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Drawing:
Mitosis
Post-Lab Questions
1. Label the arrows in the slide image below with the appropriate stage of the cell cycle.
A
C
B
F
E
D
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Mitosis
2. In what stage were most of the onion root tip cells? Based on what you know about cell cycle division, what does this imply about the life span of a cell?
3. Were there any stages of the cell cycle that you did not observe? How can you explain this using evidence from the cell cycle?
4. As a cell grows, what happens to its surface area to volume ratio? (Hint: Think of a balloon being blown up). How does this ratio change with respect to cell division?
5. What is the function of mitosis in a cell that is about to divide?
6. What would happen if mitosis were uncontrolled?
7. How accurate were your time predication for each stage of the cell cycle?
8. Discuss one observation that you found interesting while looking at the onion root tip cells.
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Mitosis
Experiment 2: Tracking Chromosomal DNA Movement through Mitosis
Although mitosis and meiosis share similarities, they are different processes and create very different results.
In this experiment, you will follow the movement of the chromosomes through mitosis to create somatic daughter cells.
Materials
2 Sets of Different Colored Pop-it® Beads (32 of each - these may be any color)
(8) 5-Holed Pop-it® Beads (used as centromeres)
Procedure
Genetic content is replicated during interphase. DNA exists as loose molecular strands called chromatin; it has not condensed to form chromosomes yet.
Sister chromatids begin coiling into chromosomes during prophase. Begin your experiment here:
1. Build a pair of replicated, homologous chromosomes. 10 beads should be used to create each individual sister chromatid (20 beads per chromosome pair). Two five-holed beads represent each centromere. To do this...
a. Start with 20 beads of one color to create your first sister chromatid pair. Five beads must be snapped together for each of the four different strands. Two strands create the first chromatid, and two strands create the second chromatid.
b. Place one five-holed bead flat on a work surface with the node positioned up. Then, two of the four strands into the bead to create an “I” shaped sister
Figure 5: Bead set-up. The blue beads reprechromatid. Repeat this step with the other two sent one pair of sister chromatids and the strands and another five-holed bead. black beads represent a second pair of sister
c. Once both sister chromatids are constructed, con- chromatids. The black and blue pair are homologous. nect them by their five-holed beads creating an “X” shape. d. Repeat this process using 20 new beads (of a different color) to create the second sister chromatid pair. See Figure 5 for reference.
2. Assemble a second pair of replicated sister chromatids; this time using 12 beads, instead of 20, per pair
(six beads per each complete sister chromatid strand).
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Mitosis
3. Repeat this process using 12 new beads (of a different color) to create the second set of sister chromatids. See Figure 6 for reference.
4. Configure the chromosomes as they would appear in each of the stages of the cell cycle (prophase, metaphase, anaphase, telophase, and cytokinesis). Diagram the images for each stage in the section titled “Cell Cycle Division: Mitosis
Beads Diagram”. Be sure to indicate the number of chromoFigure 6: Second set of replicated chromosomes present in each cell for each phase. somes. Cell Cycle Division: Mitosis Beads Diagram:
Prophase
Metaphase
Anaphase
Telophase
Cytokinesis
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Mitosis
Post-Lab Questions
1. How many chromosomes did each of your daughter cells contain?
2. Why is it important for each daughter cell to contain information identical to the parent cell?
3. How often do human skin cells divide? Why might that be? Compare this rate to how frequently human neurons divide. What do you notice?
4. Hypothesize what would happen if the sister chromatids did not split equally during anaphase of mitosis.
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Mitosis
Experiment 3: The Importance of Cell Cycle Control
Some environmental factors can cause genetic mutations which result in a lack of proper cell cycle control
(mitosis). When this happens, the possibility for uncontrolled cell growth occurs. In some instances, uncontrolled growth can lead to tumors, which are often associated with cancer, or other biological diseases.
In this experiment, you will review some of the karyotypic differences which can be observed when comparing normal, controlled cell growth and abnormal, uncontrolled cell growth. A karyotype is an image of the complete set of diploid chromosomes in a single cell.
Materials
*Computer Access
*Internet Access
*You Must Provide
Procedure:
1. Begin by constructing a hypothesis to explain what differences you might observe when comparing the karyotypes of human cells which experience normal cell cycle control versus cancerous cells (which experience abnormal, or a lack of, cell cycle control). Record your hypothesis in Post-Lab Question 1.
Note: Be sure to include what you expect to observe, and why you think you will observe these features. Think about what you know about cancerous cell growth to help construct this information
2. Go online to find some images of abnormal karyotypes, and normal karyotypes. The best results will come from search terms such as “abnormal karyotype”, “HeLa cells”, “normal karyotype”, “abnormal chromosomes”, etc. Be sure to use dependable resources which have been peer-reviewed
3. Identify at least five abnormalities in the abnormal images. Then, list and draw each image in the Data section at the end of this experiment. Do these abnormalities agree with your original hypothesis?
Hint: It may be helpful to count the number of chromosomes, count the number of pairs, compare the sizes of homologous chromosomes, look for any missing or additional genetic markers/flags, etc.
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Mitosis
Data
1.
2.
3.
4.
5.
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Mitosis
Post-Lab Questions
1. Record your hypothesis from Step 1 in the Procedure section here.
2. What do your results indicate about cell cycle control?
3. Suppose a person developed a mutation in a somatic cell which diminishes the performance of the body’s natural cell cycle control proteins. This mutation resulted in cancer, but was effectively treated with a cocktail of cancer-fighting techniques. Is it possible for this person’s future children to inherit this cancer-causing mutation? Be specific when you explain why or why not.
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Lab 12
Meiosis
Meiosis
Learning Objectives
•
Compare and contrast gametes and somatic cells including haploid and diploid
•
Explain how independent assortment, crossing over and mutations influence genetic variation
•
Apply meiosis to the development of genetic diseases
Introduction
Meiosis, also known as gametogenesis, only occurs in sexually reproducing organisms (plants and animals). In humans, meiosis generates four haploid cells in both males and females. These cells are called gametes. Male gametes are referred to as sperm cells, while female gametes are referred to as egg cells. Haploid cells only contain one copy (1n) of chromosomal content (n = 23 chromosomes in humans). Figure 1: Haploid gametes fuse during fertilization to form a diploid cell.
Haploid gametes fuse during fertilization and form a diploid cell. The newly fused cell contains two copies of each chromosome (2n = 46 chromosomes in humans); these chromosomes are now referred to as homologous chromosomes. This is unlike somatic cells which are produced through mitosis and already contain two copies of chromosomal content .
Meiosis I
Meiosis is similar to mitosis, but involves a second round of cellular division, therefore producing four daughter cells instead of two. The two rounds of cell division are called meiosis one and meiosis two. Meiosis I produces two haploid cells from the primary gamete.
The stages of meiosis I mirror the stages you learned about in mitosis except the two new cells are haploid instead of diploid. This includes prophase, metaphase, anaphase and telophase. However, because there are two rounds of meiosis, each stage also receives a “1” or a “2” after the name, depending on which round of division the cell is currently in. These stages are also indicated with Roman Numerals. You may recall that mitosis is preceded by a period called interphase during which cellular content is duplicated and DNA is proofread for accuracy. The same is true for meiosis, as an interphase period prepares the cell for successful meiotic division.
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Meiosis
The stages of meiosis I are outlined as follows:
•
Prophase I: DNA condenses into chromosomes. Centromeres attach themselves to coiled
(condensed) sister chromatids to hold their structure together. Homologous chromosomes pair up and form a chiasma. Crossing over may occur at this point. This means that homologous chromosomes may exchange and recombine regions of DNA. Crossing over creates a unique genetic fingerprint for the daughter cells, and contributes to the diversity seen in progeny.
Similar to mitotic prophase, the nuclear envelope of the parent cell breaks down in this phase as well. Pairs of centrioles, which serve as cellular anchors during division, appear at the mitotic spindle, located at the cellular poles.
•
Metaphase I: Homologous chromosomes migrate towards the metaphase plate. Microtubules grow from the centrioles and either link together or attach to each pair of homologous chromosomes at the centromeres. The orientation of each pair of sister chromatids is independent from all other pairs. This action is referred to at the Law of Independent Assortment. This means the chromosomes can “flip flop” as they line up, effectively shuffling their genetic information into new combinations, providing a second source of genetic variation.
•
Anaphase I: Microtubules attached to chromosomes. They then shorten in length, which pulls the homologous chromosomes apart (sister chromatids remain paired) and moving them toward opposite poles. Microtubules attached to other microtubules from opposite poles lengthen to elongate the cell.
•
Telophase I: One set of sister chromatids arrives at each pole. A new nuclear envelope begins to form, chromosomes are uncoiled back into chromatin, and a new nucleus is formed at each end of the cell.
Cytokinesis follows meiosis one, deepening the cellular cleavage formed in telophase one and fully dividing the two new daughter cells by engulfing each nuclei within the plasma membrane.
It is important to note that unlike mitosis, meiosis I creates haploid (not diploid) cells. This is because the homologous chromosomes during the first found of meiosis. They do not divide into the same cells. Instead, the sister chromatids remain coupled and are partitioned into the cell products. Therefore, the two daughter cells produced from meiosis one contain identical genetic content. They only contain one copy of the genome because all of the alleles are the same.
Meiosis Two
The two daughter cells produced during meiosis one immediately enter into prophase two of meiosis II. The meiosis two stages are described as follows:
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Meiosis
•
Prophase II: New spindle fibers form as the nucleus break down again. Chromosomes condense in the form of sister chromatids.
•
Metaphase II: Sister chromatids migrate towards the metaphase plate. Similar to metaphase one, microtubules attach to the centromeres.
•
Anaphase II: This time, sister chromatids (rather than homologous chromosomes) are separated as the microtubules shorten and pull them apart. The cell elongates as microtubules attached to additional microtubules from opposing poles lengthen.
•
Telophase II: 23 chromatids arrive at each pole. A new nucleus forms around each.
Cytokinesis again follows, resulting in four gametes.
Meiosis I
Prophase I
Metaphase I
Anaphase I
Telophase I
Meiosis II
Prophase II
Metaphase II
Anaphase II
Telophase II
Figure 2: Stages of Meiosis I and II.
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Meiosis
Checkpoints and Mutations
Meiotic division is complex and highly regulated. There are a series of checkpoints that a cell must pass before the next stage of division will begin. When these checkpoints function properly, mutations and errors are identified and repaired. Some of these checkpoints include verifying that ample nutrients are available for division and proofreading replicated
DNA strands. Problems which are found may result in a pause within meiosis to allow the appropriate proteins sufficient time to repair the error. If the error can’t be repaired, the cell may attempt apoptosis (cell death).
Checkpoints that are missing or do not function correctly may result in genetic mutations. Genetic mutations which occur in somatic cells are not heritable mutations. This means that the mutation will not be propagated amongst progeny. Mutations which occur in gametes are heritable mutations which risk genetic expression amongst future generations.
?
Did You Know...
There are many types of mutations. These range in scale from silent to lethal, and may affect a single nucleotide or a full chromosome. Trisomy 21, or Down’s Syndrome, is one example of a chromosomal abnormality. In this disease, three copies of chromosome
21 exist rather than the standard two copies. This type of abnormality is typically lethal, however, chromosome 21 is much smaller than many other chromosomes and thus the progeny are viable.
One mutation of particular concern is when the amount of genetic material existing in a cell changes. It is critical that a gamete contains only one copy of each chromosome found in a parent cell.
Definitions (Review):
Alleles – Alternate forms of the same gene.
Centriole – An organelle that forms the microtubules which separate chromosomes during cell division.
Centromere – The region of the chromosome where chromatids are held together. This forms the kinetochore. This is also where microtubules attach during cell division.
Chiasma – The point where crossover occurs in meiosis one.
Chromatids – One of two identical chromosomes (often called “sister chromatids”).
Chromosome – A threadlike structure consisting of chromatin that contains a long, continuous strand of DNA and its associated proteins.
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Meiosis
Crossing Over – The process, during meiosis I, by which two, paired homologous chromosomes are paired and exchange regions of DNA.
Cytokinesis – The final step of the cell cycle where the cell splits to produce two daughter cells.
Diploid – Cells that have two copies (2n) of each chromosome.
Haploid – Cells that have one copy (1n) of each chromosome.
Gamete – A cell containing a single copy of each chromosome (egg cells and sperm cells).
Gene – A sequence of DNA that codes for a specific detectable product such as a protein or RNA.
Genetic Variation – The diversity in the DNA found in otherwise similar organisms.
Homologous Chromosome – Chromosomes that contain the same genes, but potentially different alleles. Locus – The exact position of a gene on a chromosome.
Meiosis – The stage of the cell cycle when the nucleus is divided into daughter cells (gametes: sperm cells and egg cells) with a reduced number of chromosomes (from diploid to haploid).
Microtubules – Long polymer strands of protein that separate chromosomes during cell division.
Ploidy – The number (n) of copies of each chromosome contained in a cell (usually 1n or 2n in animals).
Tetrad – Four homologous chromosomes, two of each pair of sister chromatids, that line up during prophase I in meiosis.
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Meiosis
Pre-Lab Questions
1. Compare and contrast mitosis and meiosis.
2. What major event occurs during interphase?
Experiment 1: Following Chromosomal DNA Movement through Meiosis
In this experiment, you will model the movement of the chromosomes through meiosis I and II to create gametes.
Materials
2 Sets of Different Colored Pop-it® Beads (32 of each - these may be any color)
(8) 5-Holed Pop-it® Beads (used as centromeres)
Procedure
Part 1: Modeling Meiosis without Crossing Over
As prophase I begins, the replicated chromosomes coil and condense…
1. Build a pair of replicated, homologous chromosomes. 10 beads should be used to create each individual sister chromatid (20 beads per chromosome pair). The five-holed beads represent each centromere. To do this...
a. Start with 20 beads of the same color to create your first sister chromatid pair. Five beads must be snapped together for each of the four different strands. Two strands create the first chromatid, and two strands create the second chromatid with a 5holed bead at the center of each chromatid. This creates an “I” shape.
b. Connect the “I” shaped sister chromatids by the 5- Figure 3: Bead set-up. The blue beads represent one pair of sister chromatids and the holed beads to create an “X” shape. black beads represent a second pair of sister
c. Repeat this process using 20 new beads (of a differ- chromatids. The black and blue pair are hoent color) to create the second sister chromatid pair. mologous.
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Meiosis
2. Assemble a second pair of replicated sister chromatids; this time using 12 beads, instead of 20, per pair (six beads per each complete sister chromatid strand).
3. Pair up the homologous chromosome pairs created in Step
1 and 2. DO NOT SIMULATE CROSSING OVER IN THIS
TRIAL. You will simulate crossing over in Part 2.
5. Configure the chromosomes as they would appear in each of the stages of meiotic division (prophase I and II, meta- Figure 4: Second set of replicated chromophase I and II, anaphase I and II, telophase I and II, and somes. cytokinesis). 6. Diagram the corresponding images for each stage in the sections titled “Trial 1 - Meiotic Division
Beads Diagram”. Be sure to indicate the number of chromosomes present in each cell for each phase.
7. Disassemble the beads used in Part 1. You will need to recycle these beads for a second meiosis trial in Steps 8 - 13.
Trial 1 - Meiotic Beads Diagram:
Prophase I
Metaphase I
Anaphase I
Telophase I
Prophase II
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Meiosis
Metaphase II
Anaphase II
Telophase II
Cytokinesis
Part 2: Modeling Meiosis with Crossing Over
8. Build a pair of replicated, homologous chromosomes. 10 beads should be used to create each individual sister chromatid (20 beads per chromosome pair). Two five-holed beads represents the centromere. To do this...
a. Start with 20 beads of the same color to create your first sister chromatid pair. Five beads must be snapped together for each of the four different strands. Two strands create the first chromatid, and two strands create the second chromatid with a 5-holed bead at the center of each chromatid. This creates an “I” shape.
b. Connect the “I” shaped sister chromatids by the 5-holed beads to create an “X” shape.
c. Repeat this process using 20 new beads (of a different color) to create the second sister chromatid pair.
9. Assemble a second pair of replicated sister chromatids; this time using 12 beads, instead of 20, per pair
(six beads per each complete sister chromatid strand). Snap each of the four pieces into two five-holed beads to complete the set up.
10. Pair up the homologous chromosomes created in Step 8 and 9.
11. SIMULATE CROSSING OVER. To do this, bring the two homologous pairs of sister chromatids together
(creating the chiasma) and exchange an equal number of beads between the two. This will result in chromatids of the same original length, there will now be new combinations of chromatid colors.
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Meiosis
12. Configure the chromosomes as they would appear in each of the stages of meiotic division (prophase I and II, metaphase I and II, anaphase I and II, telophase I and II, and cytokinesis).
13. Diagram the corresponding images for each stage in the section titled “Trial 2 - Meiotic Division Beads
Diagram”. Be sure to indicate the number of chromosomes present in each cell for each phase. Also, indicate how the crossing over affected the genetic content in the gametes from Part1 versus Part 2.
Trial 2 - Meiotic Division Beads Diagram:
Prophase I
Metaphase I
Anaphase I
Telophase I
Prophase II
Metaphase II
Anaphase II
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Meiosis
Telophase II
Cytokinesis
Post-Lab Questions
1. In this experiment, how many chromosomes were present when meiosis I started?
2. In this experiment, how many nuclei are present at the end of meiosis II? How many chromosomes are in each?
3. What is the ploidy of the DNA at the end of meiosis I? What about at the end of meiosis II?
4. How are meiosis I and meiosis II different? List two reasons.
5. Why do you use non-sister chromatids to demonstrate crossing over?
6. What combinations of alleles could result from a crossover between BD and bd chromosomes?
7. Identify two ways that meiosis contributes to genetic recombination.
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Meiosis
8. Why is it necessary to reduce the number of chromosomes in gametes, but not in other cells?
9. Blue whales have 44 chromosomes in every cell. Determine how many chromosomes you would expect to find in the following:
Sperm Cell:
Egg Cell:
Daughter Cell from Mitosis:
Daughter Cell from Meiosis II:
10. Research and find a disease that is caused by chromosomal mutations. When does the mutation occur?
What chromosomes are affected? What are the consequences?
11. Diagram what would happen if sexual reproduction took place for four generations using diploid (2n) cells. 191
Lab 13
DNA and RNA
DNA and RNA
Learning Objectives
•
Compare and contrast the structure and function of DNA and RNA
•
Connect nucleotides, amino acids and proteins to the central dogma of the genetic code
•
Explain and perform a DNA extraction
Introduction
Long before scientists had any understanding of how humans acquire their physical traits, they knew that characteristics were passed on from generation to generation. Scientists also knew that traits were expressed as heritable proteins, but they had no idea what mechanism caused these patterns. However, they did know that there were three primary criteria which needed to be met. These criteria were:
1. It needed to carry information between generations.
2. It needed to express that information.
3. It needed to be easily replicable.
DNA Structure
Prior to the 1950’s, there was much debate over what the structure of a molecule that met all three criteria would look like. Though a number of people made significant contributions, in 1953 James Watson and Francis Crick won the Nobel Prize for their model of what we now know as DNA (deoxyribonucleic acid). The features of this model satisfied all of the necessary criteria.
DNA takes the form of what is commonly referred to as a “double helix”, and is often described as looking like a long twisted ladder with rungs
(Figure 1). The sides of the ladder consist of a sugar phosphate
“backbone” and the strand itself has directionality. In other words, like the words on this page, there is a set order in which a genetic information is read. DNA, for example, is read from the 5’ (five prime) to
3’ (three prime) end.
C
G
A
T
G
C
The rungs of the ladder carry information in a sequential series of four
A
different nucleotides (small molecules): Guanine (G), Adenine (A), ThyT mine (T) and Cytosine (C). These nucleotides pair up with each other
Figure 1: The hydrogen bonds formed via hydrogen bonding. A forms two hydrogen bonds with T, and G between adenine and thymine and forms three hydrogen bonds with C (Figure 1). No other combinations cytosine and guanine contribute to are ever made because of the chemical and electrical forces within the the double-helix structure of DNA. nucleotides. 195
DNA and RNA
Genes exists within chromatin, are stretches nucleotides which encode ribonucleic acid (RNA) and protein products. They are often described as units of heredity. Genes have specific locations on the
DNA strand and code for heritable traits (such as hair color). The human genome is comprised of 23 pairs of homologous chromosomes.
Homologous chromosomes carry the same genes but often different versions, or, alleles. Alleles are alternative forms of the same gene (e.g., brown vs. blue eyes). Somatic cells (e.g., bone, heart, skin, liver) contain two alleles (2n) for each gene. One allele is supplied by the parent sperm cell, and the second is supplied by the egg cell. ?
Did You Know...
Although “dogmas” are typically associated with a philosophy or belief system, The Central Dogma refers to the hypothesis that describes biological processes of transcription and translation. Specifically, The Central
Dogma describes:
DNA
(Transcription) mRNA During interphase of the cell cycle division, the DNA double helix
(Translation)
splits into a single helix (Figure 3). Each single helix then serves as a
Protein
template for a new strand of DNA. Neighboring nucleotides then bind to the single strand helix after which a new sugar-phosphate back- This fundamental process of reading the genetic code and developing probone is formed. teins from that code drives life.
Talk About It!
The specificity in which the nucleotides pair means the two new double helices (DNA) are identical to the original. It is the sequence of Based on what you know so far, do you think this process could function these nucleotides that are passed on from one generation to another,
DNA)?
in reverse (e.g., RNA as heritable information.
So the question remains, why is the sequence of these different four nucleotides so important? Simply put, they instruct your cells what proteins to make and how to make them via genetic sequences. This is a very important responsibility because, for example, humans and many other species are composed of proteins.
Serious health concerns or even fatality may occur if developed proteins are incorrect or product at an inappropriate concentration.
RNA Structure
Ribonucleic acid (RNA), also a nucleic acid, varies in structure and function from DNA. As the name states, RNA contains ribose while
DNA is missing the hydroxyl group (“deoxy”) from its ribose. RNA can be single-stranded or double stranded, but in most instances is singlestranded and is shorter than DNA. In RNA, the nucleotide thymine is replaced by uracil which bonds with adenine. There is more than type of RNA involved in protein synthesis: messenger (mRNA), transfer
(tRNA), and ribosomal (rRNA).
Figure 2: RNA structure.
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DNA and RNA
Transcription
For a cell to create proteins it needs to get coding information from DNA housed in the nucleus to ribosomes in the cytoplasm. To do this, a strand of messenger RNA (mRNA) is created by transcription of a gene located in the cell’s DNA. This mRNA then leaves the nucleus through nuclear pores and travels to a ribosome.
Translation
Proteins are simply chains of amino acids (small molecular building blocks) that are linked together. Twenty different amino acids are available to produce all the proteins in the body. Each amino acid is coded for by a three nucleotide sequence (codon) located on mRNA. The sequence of the amino acids determines the size of the protein and how it will fold, both factors that determine its function. Other factors such as charge and hydrophobicity (an aversion to water molecules) play a role in determining how a protein folds. Consider the following analogy:
“The Earth revolves around the sun.”
•
Figure 3: Proteins (a) are chains of amino acids asEach of the 12 different letters (codons) in the preceding sentence is sembled in ribosomes (b).
largely uninformative.
• When letters are assembled they create words, which have meaning.
• Linked words create a sentence (protein), which is then informative.
Consider a protein that is five amino acids long. Picking from the 20 available amino acids there are 520 different possible combinations (3,200,000). Even small proteins are typically several hundred amino acids long.
The number of different proteins that can ultimately be coded for by 20 amino acids is virtually limitless.
Figure 4: Gene expression; the central dogma of molecular biology.
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DNA and RNA
The next obvious question is how do the four nucleotides “code” for 20 different amino acids. Each
“letter” (codon) in the genetic code is made up of three nucleotides which codes for a specific amino acid. If we start with four possible nucleotides (A, T, G, C), how does your body make twenty different amino acids?
• If the “letter” is two nucleotides long, there are 16 possible “letters” (24) - This is not enough options.
• If the “letter” is three nucleotides long, there are 64 possible combinations (34) - more than what’s needed for twenty amino acids.
Like a sentence, the reader (a cell) needs to know where to start and where to stop (two more codons, for a total of 22). The remaining 42 possible combinations make up what is referred to as “the redundancy of the code”. In other words; Tim, Tom, Tam would all be the same person, it is simply three different spellings for his name. Each combination of three nucleotides is known as a “codon”.
Second Letter
U
C
UUU
U
UUC
UCU
Phe
UUA
UUG
Leu
First Letter
Leu
UCG
CCU
CCC
CCA
CUG
CCG
AUU
Ile
AUA
AUG
Met
GUC
UGC
Cys
C
Stop
UGA
Stop
A
UAG
Stop
UGG
Trp
G
CAU
His
CGU
CAC
Pro
CAA
CGC
Gln
U
Arg
C
CGA
A
CAG
CGG
G
ACU
AAU
AGU
U
ACC
AAC
ACA
GUU
G
UAA
Ser
U
UGU
UAC
UCC
CUA
AUC
UAU
Thr
AAA
ACG
AAG
GCU
GAU
GCC
Val
Asn
AGC
Ser
AGA
Lys
Asp
GAC
AGG
A
Arg
GGU
G
U
GGC
Ala
C
C
Gly
GUA
GCA
GAA
GUG
GCG
GAG
Glu
GGA
A
GGG
G
Figure 5: Chart showing how codons code for specific amino acids.
198
Third Letter
A
CUC
G
Tyr
UCA
CUU
C
A
DNA and RNA
Pre-Lab Questions
1. Arrange the following molecules from least to most specific with respect to the original nucleotide sequence: RNA, DNA, Amino Acid, Protein
2. Identify two structural differences between DNA and RNA.
3. Suppose you are performing an experiment in which you must use heat to denature a double helix and create two single stranded pieces. Based on what you know about nucleotide bonding, do you think the nucleotides will all denature at the same time? Use scientific reasoning to explain why.
Experiment 1: Coding
In this experiment, you will model the effects of mutations on the genetic code. Some mutations cause no structural or functional change to proteins while others can have devastating affects on an organism.
Materials
Red Beads
Yellow Beads
Blue Beads
Green Beads
Procedure
1. Using the red, blue, yellow and green beads, devise and lay out a three color code for each of the following letters (codon). For example Z = green : red : green.
In the spaces below the letter, record your “code”.
C:
___
E:
___
H:
___
I:
K:
L:
___
___
___
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DNA and RNA
M:
___
Create codons for:
O:
S:
T:
U:
___
___
___
___
Start:
Stop:
Space:
___
___
___
2. Using this code, align the beads corresponding to the appropriate letter to write the following sentence
(don’t forget start, space and stop): The mouse likes most cheese
a. How many beads did you use?
There are multiple ways your cells can read a sequence of DNA and build slightly different proteins from the same strand. We will not go through the process here, but as an illustration of this “alternate splicing”, remove codons (beads) 52 - 66 from your sentence above.
b. What does the sentence say now? (re-write the entire sentence)
Mutations are simply changes in the sequence of nucleotides. There are three ways this occurs:
1. Change a nucleotide(s)
2. Remove a nucleotide(s)
3. Add a nucleotide(s)
3. Using the sentence from exercise 1B:
a. Change the 24th bead to a different color. What does the sentence say now (re-read the entire sentence)? Does the sentence still make sense?
b. Replace the 24th bead and remove the 20th bead (remember what was there). What does the sentence say (re-read the entire sentence)? Does the sentence still make sense? If it doesn’t make sense as a sentence, are there any words that do? If so, what words still make sense?
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DNA and RNA
c. Replace the 20th bead and add one between bead numbers 50 and 51. What does the sentence say now? Does the sentence still make sense?
d. In 3.a (above) you mutated one letter. What role do you think the redundancy of the genetic code plays in this type of change?
e. Based on your observations, why do you suppose the mutations we made in 3.b and 3.c are called frame shift mutations?
f.
Which mutations do you suspect have the greatest consequence? Why?
Experiment 2: Transcription and Translation
DNA codes for all of the proteins manufactured by any organism (including you!). It is valuable, highly informative and securely protected in the nucleus of every cell. Consider the following analogy:
An architect spends months or years designing a building. Her original drawings are valuable and informative. She will not provide the original copy to everyone involved in constructing the building. Instead, she gives the electrician a copy with the information she needs to build the electrical system. She will do the same for the plumbers, the framers, the roofers and everyone else who needs to play a role to build the structure. These are subsets of the information contained in the original copy. Your cell does the same thing. The “original drawings” are contained in your DNA which is securely stored in the nucleus.
Nuclear DNA is “opened up” by an enzyme called helicase, and a subset of information is transcribed into
RNA. RNA is a single strand version of DNA, where the nucleotide uracil, replaces thymine. The copies are sent from the nucleus to the cytoplasm in the form of messenger RNA (mRNA ). Once in the cytoplasm, transfer RNA (tRNA) links to the codons and aligns the proper amino acids, based on the mRNA sequence.
Protein builders called ribosomes float around in the cytoplasm, latch onto the strand of mRNA and sequentially link the amino acids together that the tRNA has lined up for them. This construction of proteins from the mRNA is known as translation.
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DNA and RNA
Materials
Blue beads
*You Must Provide
Green beads
Red beads
In this experiment:
Yellow beads
•
Regular beads are used as nucleotides.
Pop-it® beads (8 different colors)
•
Pop-it® beads are used as amino acids.
*Pen or pencil
Procedure
1. Use a pen or pencil to write a five word sentence using no more than eight different letters in the space below. 2. Now, use the red, blue, green, and yellow beads to form “codons” (three beads) for each letter in your sentence. Then, create codons to represent the “start, “space” and stop” regions within your sentence.
Write the sentence using the beads in the space below:
3. How many beads did you use?
4. Assign one Pop-It® bead to represent each codon. You do not need to assign a Pop-It® bead for the start, stop and space regions. These will be your amino acids.
5. Connect the Pop-It® beads to build the chain of amino acids that code for your sentence (leave out the start, stop, and space regions).
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DNA and RNA
6. How many different amino acids did you use?
7. How many total amino acids did you use?
Experiment 3: DNA Extraction
Much can be learned from studying an organism’s DNA. The first step to doing this is extracting DNA from cells. In this experiment, you will isolate DNA from the cells of fruit.
Materials
(1) 10 mL Graduated Cylinder
nana, etc.)
(2) 100 mL Beakers
*Scissors
15 cm Cheesecloth
**DNA Extraction Solution
1 Resealable Bag
***Ice Cold Ethanol
1 Rubber Band (Large. Contains latex; please wear gloves when handling if you have a latex allergy).
*You Must Provide
Standing Test Tube
**Contains sodium chloride, detergent and water
Wooden Stir Stick
***For ice cold ethanol, store in the freezer 60 minutes before use.
*Fresh, Soft Fruit (e.g., Grapes, Strawberries, Ba-
Procedure
1. If you have not done so, prepare the ethanol by placing it in a freezer for approximately 60 minutes.
2. Put pieces of a soft fruit into a plastic zipper bag and mash with your fist. The amount of food should be equal to the size of approximately five grapes.
3. Use the 10 mL graduated cylinder to measure 10 mL of the DNA Extraction Solution. Transfer the solution from the cylinder to the bag with the fruit it in. Seal the bag completely.
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DNA and RNA
4. Mix well by kneading the bag for two minutes.
5. Create a filter by placing the center of the cheesecloth over the mouth of the standing test tube, pushing it into the tube about two inches, and securing the cheesecloth with a rubber band around the top of the test tube.
6. Cut a hole in the corner of the bag and filter your extraction by pouring it into the cheesecloth. You will need to keep the filtered solution which passes through the cheese cloth into the standing test tube.
7. Rinse the 10 mL graduated cylinder, and measure five mL of ice-cold ethanol. Then, while holding the standing test tube at a 45° angle, slowly transfer the ethanol into the standing test tube with the filtered solution. 8. DNA will precipitate (come out of solution) after the ethanol has been added to the solution. Let the test tube sit undisturbed for 2 - 5 minutes. You should begin to see air bubbles form at the boundary line between the ethanol and the filtered fruit solution. Bubbles will form near the top, and you will eventually see the DNA float to the top of the ethanol.
9. Gently insert the stir stick into the test tube. Slowly raise and lower the tip several times to spool and collect the DNA. If there is an insufficient amount of DNA available, it may not float to the top of the solution in a form that can be easily spooled or re-
Figure 6: DNA extraction. The color has
moved from the tube. However, the DNA will still be visible as been enhanced by dying the fruit with a white/clear clusters by gently stirring the solution and pushing substance that glows under black light. the clusters around the top.
Post-Lab Questions
1. What is the texture and consistency of the DNA?
2. Why did we use a salt in the extraction solution?
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DNA and RNA
3. Is the DNA soluble in the aqueous solution or alcohol?
4. What else might be in the ethanol/aqueous interface? How could you eliminate this?
5. Which DNA bases pair with each other? How many hydrogen bonds are shared by each pair?
6. How is information to make proteins passed on through generations?
7. Watch the following Virtual Lab demonstrating DNA Extraction. In this experiment, how do the Lysis Solution and the Salt Solution vary by function?
8. Identify one step which was included in the Virtual Lab which was not required in the hands-on experiment. Then, identify one step which was included in the hands-on experiment, but not the virtual lab. Why weren’t these steps required?
205
Lab 14
Mendelian Genetics
Mendelian Genetics
Learning Objectives
•
Explain how Mendel’s work formed the foundation of modern genetics including the law of segregation and the law of independent assortment
•
Relate genes to homozygous, heterozygous, dominant alleles, recessive alleles, genotype and phenotype •
Use monohybrid and dihybrid crosses to analyze patterns of inheritance including dominance, incomplete dominance and co-dominance
Introduction
In 1866, Gregor Mendel (an Austrian Monk), published a paper entitled “Experiments in Plant Hybridization”.
It went largely unnoticed until 1900 when it was “rediscovered” and subsequently became the basis for what we now refer to as Mendelian Genetics. Mendel was the first to recognize:
•
Inherited characteristics are determined by specific factors (now recognized as genes).
•
These factors occur in pairs (homologous genes).
•
When gametes form, these factors segregate so that each gamete contains only one.
The Law of Independent Assortment
These original observations lead to what we now refer to as
The law of segregation and the law of independent assortment. The law of segregation states that during meiosis, homologous (paired) chromosomes split. The law of independent assortment states that during meiosis, each homologous chromosome has an equal chance of ending up in either gamete, and alleles for individual genes segregate with the chromosomes on which they are located (Figure 1).The law of independent assortment allows for genetic recombination. The following equation can be used to determine the total number of Figure 1: Law of Independent Assortment. possible genotype combinations for any particular number of genes: 2g= Number of possible genotype combinations (where “g” is the number of genes)
•
1 gene: 21= 2 genotypes
•
2 genes:22= 4 genotypes
•
3 genes: 23 = 8 genotypes
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Mendelian Genetics
Genotype and Phenotype
Genotype refers to the combination of alleles for a particular trait. Phenotype refers to the appearance of that combination of alleles. In other words, genotype determines phenotype. Genotype cannot be seen and is represented by letters. Using corn as an example (Figure 2):
•
The large chromosome has the gene for kernel color (Y = yellow, y = blue).
•
The small chromosome has the gene for kernel texture (S = smooth (green); s = wrinkled (red)).
Dominance
In our example, the genotype of the diploid cell is Yy, Ss, while the phenotype is Yellow and Smooth. When a dominant allele is present, that characteristic is expressed, regardless of the second allele. In this case both the Yy and
YY offspring will be yellow. The phenotype inscribed by a recessive allele is only expressed when both alleles are recessive. In this case, only the yy combination is blue. The dominant allele is always represented by capital letters, while the recessive is represented by lower case letters. Any letter Figure 2: If one dominant allele is present the can be used to represent a gene, but usually the first letter of dominant phenotype is expressed. If two recessive alleles combine a recessive phenotype is the dominant phenotype used (ex., Yellow). expressed. Homozygous and Heterozygous Genotypes
When both alleles of a gene are the same they are said to be homozygous. If they are different they are said to be heterozygous. When gametes form, these factors segregate so that each gamete contains only one allele for each gene. Remember, alleles are one copy of a gene and are coded within a chromosome.
Incomplete Dominance and Co-Dominance
Alleles can exhibit incomplete dominance and co-dominance. An example of incomplete dominance is the cross of two plants, one with red flowers and one with white, whose offspring have pink flowers. In the case of co-dominance, the same cross would result in red and white striped flowers.
Predicting Inheritance
If we know the genotype of two parents we can predict both the genotype and phenotype of their offspring using a Punnett Square. A monohybrid cross is a cross between two parents (P), looking at a single gene. In this example, both parents are pure breeding (homozygous); one for the yellow color and one for the blue color. This cross can be shown as a Punnett Square (Figure 3), with each parent (P) contributing a single gamete. The offspring (F1) are determined by adding the gamete of each parent (P) (Row and Column). The cross of the (F1) generation, known as the F2 generation, is shown in Figure 4.
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Mendelian Genetics
When the P generation is crossed one hundred percent offspring are heterozygous (Y , y) and yellow in appearance even though one parent had blue kernels (Figure 3). When two offspring from the F1 generation are crossed their offspring have a 75% chance of having yellow kernels and 25% chance of having blue kernels (Figure 4). While the recessive phenotype disappears in the F1 generation it reappears in the F2 generation. Parent 1
Y
Y
y
Y y
Y y
y
Y y
Y y
Parent
2
Figure 3: Punnett Square Monohybrid Cross F1.
The Punnett square can also predict the F1 for multiple genes. Using our corn example, let’s look at two genes (color and texture), also known as a dihybrid cross. In this example we use two parents that are heterozygous for both traits (Figure 5), using the gametes we already identified in (Figure 3).
Parent 1
Y
y
Y
Y Y
Y y
y
Y y
y y
Parent
2
Figure 4: Punnett Square Monohybrid Cross F2.
The F2 phenotypes are:
Yellow & Smooth: 9
Yellow & Wrinkled: 3
Blue & Smooth: 3
Blue & Wrinkled: 1
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Mendelian Genetics
Y s
Y S
y S
y s
Y s
Y Y s s
Y Y S s
y Y S s
y Y s s
Y S
Y Y s S
Y Y S S
y Y S S
y Y s S
y S
Y y s S
Y y SS
y y S S
y y s S
y s
Y y s s
Y y S s
y y S s
y y s s
Figure 5: Punnett square of a dihybrid cross (F1).
Pre-Lab Questions
1. In a species of mice, brown fur color is dominant to white fur color. When a brown mouse is crossed with a white mouse all of their offspring have brown fur. Why did none of the offspring have white fur?
2. Can a person’s genotype be determine by their phenotype? Why or why not?
3. Are incomplete dominant and co-dominant patterns of inheritance found in human traits? If yes, give examples of each.
4. Consider the following genotype: Yy Ss Hh. We have now added the gene for height: Tall (H) or Short (h).
a. How many different gamete combinations can be produced?
b. Many traits (phenotypes), like eye color, are controlled by multiple genes. If eye color were controlled by the number of genes indicated below, how many possible genotype combinations would there be in the following scenarios?
5 Eye Color Genes:
10 Eye Color Genes:
20 Eye Color Genes:
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Mendelian Genetics
Experiment 1: Punnett Square Crosses
In this experiment you will use monohybrid and dihybrid crosses to predict patterns of inheritance.
Materials
Blue Beads
Yellow Beads
Green Beads
(2) 100 mL Beakers
Red Beads
Permanent Marker
Procedure
Part 1: Punnett Squares
1. Set up and complete Punnett squares for each of the following crosses: (remember Y = yellow, and y = blue) Y Y and Y y
Y Y and y y
2. What are the resulting phenotypes?
3. Are there any blue kernels? How can you tell?
4. Set up and complete a Punnett squares for a cross of two of the F1 from Step 1 (above).
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Mendelian Genetics
5. What are the genotypes of the F2 generation?
6. What are their phenotypes?
7. Are there more or less blue kernels than in the F1 generation?
8. Identify the four possible gametes produced by the following individuals:
a) YY Ss:
______
______
______
______
b) Yy Ss:
______
______
______
______
c) Create a Punnett square using these gametes as P and determine the genotypes of the F1:
What are the phenotypes? What is the ratio of those phenotypes?
Part 2 and 3 Setup
1. Use the permanent marker to label the two 100 mL beakers as “1” and “2”.
2. Pour 50 of the blue beads and 50 of the yellow beads into Beaker 1. Sift or stir the beads around to create a homogenous mixture.
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Mendelian Genetics
3. Pour 50 of the red beads and 50 of the green beads into Beaker 2. Sift or stir the beads around to create a homogenous mixture.
Assumptions for the remainder of the experiment:
•
Beaker 1 contains beads that are either yellow or blue.
•
Beaker 2 contains beads that are either green or red.
•
Both beakers contain approximately the same number of each colored bead.
•
These colors correspond to the following traits (remember that Y/y is for kernel color and S/s is for smooth/wrinkled):
1. Yellow (Y) vs. Blue (y)
2. Green (G) vs. Red (g).
Part 2: Monohybrid Cross
1. Randomly (without looking) take two beads out of Beaker 1. This is the genotype of Individual #1. Record the genotype in Table 1. Do not put these beads back into the beaker.
Table 1: Parent Genotypes: Monohybrid Crosses
Genotype of In-
Genotype of Indi-
P1
P2
P3
P4
Generation
P
2. Repeat Step 1 for Individual #2. These two genotypes represent the parents (generation P) for the next generation. 3. Set up a Punnett square and determine the genotypes and phenotypes for this cross. Record your data in Table 2
4. Repeat Step 3 four more times (for a total of five subsequent generations). Return the beads to their respective beakers when finished.
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Mendelian Genetics
Table 2: Generation Data Produced by Monohybrid Crosses
Parents
Possible Offspring Genotypes
Possible Offspring Phenotypes
Genotype
Ratio
Phenotype
Ratio
P
P1
P2
P3
P4
Part 3: Dihybrid Cross
1. Randomly (without looking) remove two beads from of Beaker 1 and two beads from Beaker 2. These four beads represent the genotype of Individual #1. Record this information in Table 3
2. Repeat Step 1 to obtain the genotype of Individual #2. Record the phenotypes of both individuals in Table 3.
Table 3: Parent Genotypes: Dihybrid Crosses
Genotype of In-
Genotype of Indi-
P1
P2
P3
P4
Generation
P
3. Determine what the possible genotypes might be if each individual produced gametes. Record these possible genotypes in Table 4.
4. Predict the ratio of possible genotypes which could be produced by a cross between Individual #1 and
Individual #2? Record your predictions in Table 4.
Hint: Think back to the example the dihybrid cross in the Introduction. Record your prediction in Table
2.
5. Set up a Punnett square and determine the genotypes and phenotypes for this cross.
6. Repeat Step 5 four additional times (for a total of five subsequent generations).
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Mendelian Genetics
Table 4: Generation Data Produced by Dihybrid Crosses
Parents
Possible Offspring Genotypes
Possible Offspring Phenotypes
Genotype
Ratio
Phenotype
Ratio
P
P1
P2
P3
P4
Post-Lab Questions
Part 2: Monohybrid Cross
1. How much genotypic variation do you find in the randomly picked parents of your crosses?
2. How much in the offspring?
3. How much phenotypic variation?
4. Is the ratio of observed phenotypes the same as the ratio of predicted phenotypes? Why or why not?
5. Pool all of the offspring from your five replicates. How much phenotypic variation do you find?
6. What is the difference between genes and alleles?
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Mendelian Genetics
7. How might protein synthesis execute differently if a mutation occurs?
8. Organisms heterozygous for a recessive trait are often called carriers of that trait. What does that mean?
9. In peas, green pods (G) are dominant over yellow pods. If a homozygous dominant plant is crossed with a homozygous recessive plant, what will be the phenotype of the F1 generation? If two plants from the F1 generation are crossed, what will the phenotype of their offspring be?
Part 3: Dihybrid Cross
1. How similar are the observed phenotypes in each replicate?
2. How similar are they if you pool your data from each of the five replicates?
3. Is it closer or further from your prediction?
4. Did the results from the monohybrid or dihybrid cross most closely match your predicted ratio of phenotypes?
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Mendelian Genetics
5. Based on these results; what would you expect if you were looking at a cross of 5, 10, 20 independently sorted genes?
6. Why is it so expensive to produce a hybrid plant seed?
7. In certain bacteria, an oval shape (O) is dominant over round (o) and thick cell walls (T) are dominant over thin (t). Show a cross between a heterozygous oval, thick cell walled bacteria with a round, thin cell walled bacteria. What are the phenotype of the F1 and F2 offspring?
219
Lab 15
Population Genetics
Population Genetics
Learning Objectives
•
Explain how genetic drift, the founder effect, mutations and natural selection affect a population’s gene pool
•
Use the Hardy-Weinberg equation to calculate gene frequencies within a population
•
Analyze the effects of stochastic events on genetic variation and frequency in a gene pool
Introduction
In the previous lab we looked at how genes are passed on to offspring. In this lab, the exercises are designed to look at individual genes (two alleles, one dominant, one recessive). However, we will be looking at their presence, prevalence and distribution at the population level.
Gene Frequency
The gene pool is the sum of all genes and their corresponding alleles in a given population. Take a look at the population of 100 brown and white mice in Figure 1. The color brown (B) is dominant. The standard for naming alleles is to use the case of the dominant trait, with the lower case to represent the recessive allele.
Their gene pool is B, b.
Gene frequency refers to how many times each allele is found in the population. These 100 mice have 200 alleles: •
55 heterozygous mice (B, b) have 55 B alleles and 55 b alleles.
•
27 homozygous recessive mice (b, b) have 54 b alleles (2 x 27 “b”= 54).
•
18 homozygous dominant mice (B, B) have 36 B alleles (18 x 2 “B” = 36).
The gene frequency of the population is:
B: 91 b: 109
Figure 1: Mouse population.
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Population Genetics
Often this is represented as a percentage of the dominant gene. In this case, the percentage of B is 45.5%
(=91/200). Note that the dominant gene is less prevalent than the recessive gene. This is not unusual. It is important to remember that dominance has no direct relationship to prevalence.
Hardy-Weinberg Equilibrium
To estimate the frequency of alleles in a population, we can use the Hardy-Weinberg equation. According to this equation: p = the frequency of the dominant allele (represented here by A) q = the frequency of the recessive allele (represented here by a)
For a population in genetic equilibrium:
p + q = 1.0 (The sum of the frequencies of both alleles is 100%.)
(p + q)2 = 1
So
p2 + 2pq + q2 = 1
The three terms of this binomial expansion indicate the frequencies of the three genotypes:
p2 = frequency of AA (homozygous dominant)
2pq = frequency of Aa (heterozygous) q2 = frequency of aa (homozygous recessive)
Stochastic Events
Many stochastic events change both the gene pool and gene frequencies over time. These parameters can also change as a result of mutation and natural selection.
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Population Genetics
Mutations are a change in the sequence of DNA. Most mutations do not change the phenotype and confer no advantages or disadvantages to the individual. Each of us has hundreds and probably thousands of mutations.
Nonetheless, some mutations do change the protein coded for by a gene. The vast majority of these mutations are lethal and the embryo never fully develops. Occasionally mutations do not effect embryonic development and the offspring is born without complication.
Pre-Lab Questions
Assumptions:
•
There are approximately 3,000,000,000 base pairs in the mammalian genome (genes constitute only a portion of this total).
•
There are approximately 10,000 genes in the mammalian genome.
•
A single gene averages 10,000 base pairs in size.
•
Only 1 out of 3 mutations that occur in a gene result in a change to the protein structure.
In the mammalian genome:
1. How many total base-pairs are in all the mammalian genes?
2. What proportion (%) of the total genome does this represent?
3. What is the probability that a random mutation will occur in any given gene?
4. What is the probability that a random mutation will change the structure of a protein
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Population Genetics
Note: In the following experiments on gene pool, gene frequency, and genetic diversity; assume there are four alleles for color and that they are all homologous.
Experiment 1: Genetic Variation
Genetic variation is simply the genetic difference within or between populations, in the gene pool and/or gene frequency.
Consider the following two populations of butterflies (Figure 2):
Figure 2: Butterfly populations.
Assumptions: Both populations contain the same four colors of butterflies, thus the gene pool is the same.
However, the distribution of colors within that population is different, thus their gene frequencies are different.
Materials
Blue Beads
Yellow Beads
Red Beads
(2) 100 mL Beakers
Green Beads
(2) 250 mL Beakers
Procedure
1. Pour 50 blue beads and 50 red beads into a 250 ml beaker. Without looking, randomly take 50 beads from the 250 mL beaker and place them in a 100 mL beaker (this is Beaker #1).
2. Pour 50 green beads and 50 yellow beads into a second 250 mL beaker. Without looking, randomly take
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Population Genetics
20 beads from the 250 mL beaker and place them in the other 100 mL beaker (this is beaker #2).
Note: When done, return beads to their respective beakers (1 or 2) for use in the next experiment.
Post-Lab Questions
1. What is the gene pool of beaker #1?
2. What is the gene pool of beaker #2?
3.
What is the gene frequency of beaker #1?
4.
What is the gene frequency of beaker #2?
5. What can you say about the genetic variation between these populations?
Experiment 2: Genetic Drift
Genetic drift, the variation of the gene pool and/or gene frequency of a population, can result from a variety of stochastic (random) means. Consider the following population of butterflies who have half of their habitat destroyed by wildfire (Figure 3):
Figure 3: Butterfly populations before and after wildfire.
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Population Genetics
The remaining population has 50% of the initial gene pool (two colors) and the gene frequency is different. As these individuals reproduce, their offspring will no longer reflect the original population.
Materials
Blue Beads
Beads in Beaker #1 (from Experiment 1)
Green Beads
Beads in Beaker #2 (from Experiment 1)
Red Beads
(1) 100 mL Beaker
Yellow Beads
(1) 250 mL Beaker
Procedure
1. From the 250 mL beaker containing green and yellow beads, take 10 beads and place them into the unused 100 mL beaker (this is Beaker #3).
2. Randomly remove half of the beads from Beakers #1, #2, and #3. These are the individuals that survived the fire. Keep them separated so they can be returned to their proper beaker. You will use them again in the next lab.
3. Record your results and place the beads back in their respective beakers.
4. Repeat this process four more times (five total).
Post-Lab Questions
1. What observations can you make regarding the gene pool and gene frequency of the surviving individuals?
2. Do the results vary between the populations represented by beakers #1, #2 and #3? Why or why not?
3. What observations can you make about the genetic variation between the parent and surviving populations?
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Population Genetics
Experiment 3: Stochastic Events
Consider the same population of butterflies is in the path of a hurricane. All survive, but 10 are blown to a new location. These 10 start a new population; their progeny will reflect the founder’s gene pool. This is known as the founder effect.
Figure 4: Butterfly populations before and after hurricane.
Materials
Beaker #1 (from Experiment 1)
Beaker #2 (from Experiment 1)
Beaker #3 (from Experiment 2)
Procedure
1. Remove 10 individuals from Beaker #1, five from Beaker #2 and two from Beaker #3. These are the founders of your new population.
2. Record your results and place the beads back in their respective beakers.
3. Repeat this process four more times (five total).
Note: When you are finished with this experiment, return the beads to their appropriate bags.
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Population Genetics
Post-Lab Questions
1. What observations can you make regarding the gene pool and gene frequency of the founding individuals?
2. Do these results vary between the populations founded by beakers #1, #2 and #3? Why or why not?
3. What observations can you make about the genetic variation between the parent and founding populations?
4. Suppose you have a population of 300 butterflies. If the population experiences a net growth of 12% in the following year, how many butterflies do you have?
5. Now suppose you have 300 eggs, but only 70% of those eggs progress to become a caterpillar, and only 80% of the caterpillar progress to become an adult butterfly. How many butterflies do you have?
(For simplicity, assume that all butterflies survive to the next year, in this example)
6. Suppose you have a population of 150 butterflies, but a wildfire devastates the population and only 24 butterflies survive. What percent does the colony decrease by?
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Population Genetics
Experiment 4: Natural Selection
Natural selection is a selection pressure that effects phenotypes in one of three ways:
1. It will create an adaptive advantage.
2. It will create an adaptive disadvantage.
3. It will remain entirely neutral.
A classic example to illustrate natural selection comes from England. Prior to the Industrial Revolution, the native moths were normally a light color, though darker versions of the same species existed. The lighter color blended with the light bark of the local trees, while the darker moths experienced a higher predation rate - they were easier for birds to spot and fewer survived to reproduce. As England entered the Industrial
Revolution they began burning fossil fuels with little regard to the pollutants they were emitting. The trunks of the trees became coated with soot and their color darkened. The lighter moths became more conspicuous and the darker moths were better camouflaged. The proportion of white to dark moths changed.
Materials
(1) 100 mL Beaker
Red Beads
Blue Beads
Access to a Printer
Procedure
1. Print the two sheets of paper marked Blue Habitat (Figure 5) and Red Habitat (Figure 6), from your manual (found at the end of this procedure).
2. Place 50 red and 50 blue beads into a 100 mL beaker.
3. Mix them well and pour them onto the sheet marked Red Habitat.
4. Keep the beads that fall onto habitat that matches their color.
5. For each bead that you keep (and return to the beaker), add another bead of the same color to the beaker (discard the rest).
6. Repeat this three times.
7. Record the remaining colors.
Blue _____________ Red _____________
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Population Genetics
Do you observe a selective advantage for the red or blue beads? Why?
8. Repeat the process using the Blue Habitat with the remaining beads.
What beads remain now?
Blue _____________
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Red _____________
Population Genetics
Figure 5: Red Habitat
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Population Genetics
Figure 6: Blue Habitat
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Population Genetics
Post-Lab Questions
1. How did the distribution of phenotypes change over time?
2. Is there a selective advantage or disadvantage for the red and/or blue phenotypes?
3. What phenotypic results would you predict if starting with the following population sizes?
A. 1000:
B. 100:
C. 10:
4. Assume that you live in a country with 85 million people that consistently experiences an annual growth rate of 4.2%. If this population continues to grow at the same rate for the next 5 years, how many people will live in the country (round to the nearest whole number).
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Population Genetics
Experiment 5: Sickle Cell Anemia Inheritance Patterns
Sickle cell anemia is a genetic disease (one base pair mutation that changes a protein). It is more common in those of African ancestry. “S” will represent the normal dominant allele and “s” the recessive sickle allele.
They are co-dominant alleles – SS is normal, Ss is not fatal, ss is debilitating, painful and often fatal.
Materials
(1) 100 mL Beaker
Blue Beads
Red Beads
Procedure
1. Place 25 red (S) and 25 blue (s) beads into the 100 mL beaker and mix well.
2. Randomly (without looking) remove two beads. Repeat 10 times (without returning the beads to the beaker), each time recording if it was a SS, Ss or ss.
3. Assume that each individual with the ss genotype passed away, and remove ss instances from the population.
4. The remaining beads survived and reproduced.
5. Count how many red and blue beads remained (separately) and place twice that number back in the beaker. 6. Repeat the process seven times.
Post-Lab Questions
1. What is the remaining ratio of alleles?
2. Have any been selected against?
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Population Genetics
3. Given enough generations, would you expect one of these alleles to completely disappear from the population? Why or why not?
4. Would this be different if you started with a larger population? Smaller?
5. After hundreds or even thousands of generations both alleles are still common in those of African ancestry. How would you explain this?
6. The worldwide distribution of sickle gene matches very closely to the worldwide distribution of malaria
(Figure 7). What is the significance of this?
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Population Genetics
238
Figure 7: Distribution of Malaria (top) and Sickle Cell trait (bottom).
Lab 16
Taxonomy
Taxonomy
Learning Objectives
•
Apply Linnaean’s classification system to analyze relatedness
•
Explain how advances in taxonomy such as phylogenetics have changed the way organisms are classified •
Compare and contrast binomial nomenclature and common name
Introduction
Taxonomy is the classification (organization) of organisms into related groups. Very early on, scientists recognized the importance of classifying or ranking organisms. In the 1700s, Carl Linnaeus (also known as Carl von Linné or Carolus Linnaeus) developed what is now known as the Linnaean Classification System. At the highest level in this system, everything in nature was divided into three kingdoms: mineral, vegetable, and animal. Linnaeus then ranked sequentially smaller groups within each kingdom as class, order, genus, species and variety. This contribution to science earned him the title “The Father of Taxonomy”. His system is still, in an evolved form, used today.
Starting with domain, there is a continual increase in specificity as organisms are classified into smaller and smaller categories (Figure 1 ). In other words, the categories get smaller in terms of the number of organisms that are included. As illustrated in Figure 1, the Linnaean system classifies organisms into sequential groups:
•
Domain
•
Kingdom
•
Phylum
•
Class
•
Order
•
Family
•
Genus
•
Species
Interestingly, there is not a consensus definition of
‘species’ among scientists at this time. The biological species concept is the most commonly used definition. Basically it defines a species as a group of individuals that can interbreed and create viable offspring. Sub-species are used in some classifications and is generally accepted, but not always included in the Linnaean system.
Figure 1: Taxonomy Tree. To use this tree, one would begin at the bottom and work up.
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Taxonomy
Table 1: Sample Classifications of Humans and a Red Maple Tree
Example
Human Being
Red Maple
Domain
Eukarya
Eukarya
Kingdom
Animalia
Plantae
Phylum
Chordata
Tracheophyta
Class
Mammalia
Angiospermae
Order
Primates
Sapindales
Family
Hominidae
Acerceae
Genus
Homo
Ace
Species
Sapien
Rubrum
A useful tool to remembering the order of the Linnaean classification system is developed by creating a mnemonic phrase using the first letter of each classification. For example, Daring Kids Pick Cauliflower
Over Fresh Grown Strawberries. Can you create any mnemonics to help remember the Linnaean classification system?
Binomial Nomenclature
Carl Linnaeus also helped to standardize the naming of species by introducing the binomial (two name) species name. Binomial nomenclature uses the genus and species as the formal name of all organisms because they are the two most specific levels therefore unique to the organism. For example, human beings are Homo sapiens. Note that the genus is capitalized while species is not and, since these are Latin names, they are italicized. Some organisms can have more than one common name (i.e. cougar, puma and mountain lion). Using binomial nomenclature specifies a species.
Advances in Taxonomy
Taxonomy is in a constant state of flux. Traditional groupings of organisms were dependent on mostly morphological traits, physiological similarities and, to some extent, embryological events. Consider what techniques were available in the 1700s when Carl Linnaeus described his system for ranking life. Gross anatomy could be studied and identified, but most forms of microscopy were not available.
Phylogenetics
With advances in molecular techniques, nucleic acid (DNA) sequences are used to classify organisms based on their evolutionary history. These advancements have given birth to the field of phylogenetics.
Phylogenetics is the field of study that determines the evolutionary relationships among groups of organisms using morphological traits, behavioral information and molecular data. Ribosomal RNA (rRNA) is cur-
242
Taxonomy rently one of the genes of choice for phylogenetic studies. rRNA is necessary for the production of proteins and, because all life forms require proteins, it follows that all organisms include rRNA genes. Sequences from specific regions of an rRNA gene demonstrate similarities amongst different organisms, enabling evolutionary relationships to be explored. It is well accepted that the more similar the sequences are between two organisms, the more closely related the organisms. As DNA sequencing techniques improve (and costs decrease) other regions of the genome are also being sequenced and compared.
Hot Topics in Taxonomy
Keep in mind that new techniques aid in classifications and, at the same time, cause previously well accepted classifications to become obsolete. Currently, many sources list three domains: Bacteria, Archaea, and Eukarya. Within Eukarya are four kingdoms: Protista, Plantae, Fungi, and Animalia. The Kingdom Protista has historically been a ‘catch-all’ category for all single-celled eukaryotic organisms; but, not plants, fungi or animals. However, molecular data has revealed distinct relationships within the eukaryotes that has led to suggestions for new kingdoms and “supergroups”. In this proposal, organisms that share a common ancestor are grouped together and organisms that were previously placed in the Kingdom Protista are distributed throughout the new categories. Recall that the organization of organisms based on shared common ancestors is considered phylogeny. The recently suggested revised “supergroups” are Bacteria, Archaea, Excavata, Amoebozoa, Opisthokonta, Rhizaria, Chromalveolata, and Archaeplastida. In this organizational system, animals fall within Opisthokonta and plants are placed in Archaeplastida.
Classifications will continue to evolve as scientists are able to delve at increasingly deeper molecular levels. For now, the Kingdom Protista will continue to be used in this manual as we briefly survey the three domains (Bacteria, Archaea and Eukarya) and four kingdoms (Protista, Plantae, Fungi, and Animalia).
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Taxonomy
Pre-Lab Questions
1. Use the following classifications to determine which organism is least related out of the three. Explain your rationale.
Table 2: Classifications
Classification Level
American Green Tree
Frog
European FireBellied Toad
Eastern Newt
Domain
Eukarya
Eukarya
Eukarya
Kingdom
Animalia
Animalia
Animalia
Phylum
Chordata
Chordata
Chordata
Class
Amphibia
Amphibia
Amphibia
Order
Anura
Anura
Caudata
Family
Hylidae
Bombinatoridae
Salamandridae
Genus
Tursipops
Bombina
Notophthalmus
Species
cinerea
bombina
viridescens
2. How has DNA sequencing affected the science of classifying organisms?
3. You are on vacation and see an organism that you do not recognize. Discuss what possible steps you can take to classify it.
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Taxonomy
Experiment 1: Dichotomous Key Practice
In this experiment, you will identify organisms by their binomial nomenclature using a dichotomous key. A dichotomous key is an identification tool that starts with a broad defining characteristics and splits into two options until an organism can be identified.
Materials
Images of Organisms (Figure 2)
Dichotomous Key (Figure 3)
Procedure
1. Start by observing organism i (Figure 2). Once you have taken notice of its physical characteristics, use the dichotomous key (Figure 3) to identify the organism.
2. Start at number 1 on the key and decide if the organism has feature “1a” or feature “1b”.
3. Which ever option you choose, follow the dotted line over to the next step that will state “Go to #”.
4. Go to that number and again decide between the two options.
5. Eventually you will wind up at a two name scientific name.
6. Once you have identified organism i record your finding in Table 3.
7. Repeat steps 1-6 for all of the organisms in the table.
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Taxonomy
i.
ii.
iii.
iv.
vi.
v.
vii.
viii. ix. x.
xi.
xiii. xii. 246
Figure 2: Organisms to be identified.
Taxonomy
1a. Animal…………………………………………………………………………………..………………………………………………………………………...……………...Go to 5
1b. Not an animal…………………………………………………………………….…………………………………………………………………………….…………….Go to 2
2a. Plant that bares fruit or nut………………………………………………….……………………………………………………………………..…………………..Go to 3
2b. Plant that bares only flowers……………………………………………….……………………………………………………………………...……Lonicera iaponica
3a. Plant bares nut………………………………………………………………...……………………………………………………………………..…………….…Quercus abla
3b. Plant bares fruit………………………………………………………………..………………………………………………………………………..…………………..Go to 4
4a. Plant bares yellow fruit when ripe………………………………………….……………………………………………………………….…………Musa acuminate
4b. Plant bares red fruit when ripe……………………………………………..………………….……………………………………...…………Vaccinium Oxycoccus
5a. Vertebrate (backbone)………………………………………………………….……………………………………………..………………………………………….Go to 8
5b. Invertebrate (no backbone)………………………………………………….……………………………………………….………………………………………..Go to 6
6a. Exoskeleton (hard outer covering)……………………………………………………………………………………...…………………………………………..Go to 7
6b. So flesh with shell……………………………………………………………….…………………………………………………...………….………………Helix aspersa
7a. Eight legs……………………………………………………………………………….……………………………………………..………………...….………Euathlus smithi
7b. Six legs……………………………………………………………………………….……………………………………………………....…….……………Oryctes nasicornis
8a. Has fur………………………………………………………………………………..………………………………………………………..………………………Mus musculus
8b. Has feathers……………………………………………………………………….………………………………………..………………………………………………...Go to 9
9a. Long sharp beak…………………………………………………………………….………………………………………….…………………………………………..Go to 10
9b. Short triangular beak…………………………………………………………...…………………………………………….…………………………………………Go to 12
10a. Thick, mul color beak………………………………………………………...…………………………………………………………... ……Ramphastos vitellinus
10b. Thin, needle‐like beak………………………………………………………...………………….…………………………………………………………………..Go to 11
11a. Red colora on on crown……………………………………………..………………………………………………………………………………Dryocopus pileatus 11b. Green colora on on tail………………………………………………….……………………………………………..……………………..Selasphorus platycercus
12a. Gray and orange colora on………………………………………………..……………………………………………..………………………Taeniopygia guƩata
12b. Yellow body colora on……………………………………………………….…………………………………...……………………………..…………Carduelis trisƟs
Figure 3: Dichotomous key.
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Taxonomy
Table 3: Dichotomous Key Results
Organism
i ii iii iv v vi vii viii ix x xi xii xiii
248
Binomial Name
Taxonomy
Post-Lab Questions
1. What do you notice about the options of each step as they go from number one up.
2. How does your answer from question one relate to the Linnaean classification system?
Experiment 2: Classification of Organisms
Materials
Table 2
Figure 4
Procedure
1. Select the first organism from Table 4 (E. coli).
2. Use the “tree” (Figure 4; located at the end of this procedure) start at the base, and answer each question until the organism reaches the end of a “branch”. Write the organisms name in the red box.
3. Repeat this for the remaining organisms.
4. After classification, fill in Table 2 with the correct kingdom for each organism.
Table 4: Key Characteristics of Some Organisms
Organism
Kingdom
Defined Nucleus
E. Coli
Mobile
Cell Wall
Yes
Yes
Yes
No
Yes
Protozoa
Yes
Mushroom
Yes
Sunflower
Yes
No
Bear
Yes
Yes
Photosynthesis
Unicellular
Yes
Yes
Yes
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Taxonomy
Start
Does the organism have a defined nucleus?
Yes
No
Kingdom:
Bacteria
Is the organism mobile?
Yes
Kingdom:
No
No
Fungi
Yes
Kingdom:
Does the organism perform photosynthesis?
Does the organism have specialized cells? Yes
Plant
No
Kingdom:
Kingdom:
Animal
Protist
Figure 4: Experiment 2 - Classification of Organisms Flow Chart
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Taxonomy
Post-Lab Questions
1. Did this series of questions correctly organize each organism? Why or why not?
2. What additional questions would you ask to further categorize the items within the kingdoms (Hint: think about other organisms in the kingdom and what makes them different than the examples used here)?
3. Do you feel that the questions asked were appropriate? What questions would you have asked?
251
Lab 17
Bacteria and Archaea
Norris Geyser Basin in Yellowstone Na onal Park
Bacteria and Archaea
Concepts to Explore
•
Identify characteristics of prokaryotes including morphology and energy production
•
Distinguish between prokaryotes that are autotrophic, heterotrophic and chemotrophic
•
Distinguish between prokaryotes based on their morphology including cocci, bacilli, and spirilla
•
Describe the structure of bacterial DNA
Introduction
Bacteria are simple organisms only microns in length, are found everywhere on the planet and are some of the first organisms found in the fossil record. Their varied habitats include soil, acidic hot springs, radioactive waste, seawater, glacial ice and deep in the Earth’s crust.
Bacteria are prokaryotes, cells with no organized nucleus or organelles. Their
DNA is not condensed into chromosomes, but is found as circular loops that float around the cytoplasm. It has been proposed that prokaryotes evolved from endosymbiotic associations after a larger bacterium engulfed a smaller one. This may also explain how mitochondria, chloroplasts, and other organelles developed.
?
Did You Know...
Halobacteria is a group of organisms which actually belongs to the archaea domain (not bacteria). Surprisingly, they thrive in concentrated brine nine
Most bacteria are heterotrophic, meaning they cannot manufacture their own times the salinity of sea food. Often they excrete digestive enzymes that decompose the environment water, and remain viable in around them and use the byproducts as energy. However, some are Auto- dry salt crystals for years.
trophic, either obtaining energy from sunlight through photosynthesis, or
Chemoautotrophic, creating energy from inorganic chemicals.
Some prokaryotes have whip-like flagella for locomotion, while others have hair-like pili that allow them to adhere to surfaces. Many bacteria are surrounded by an exterior capsule (coating), which prevents white blood cells (the body’s defense cells) from identifying and destroying them. This coat also makes them stick to target surfaces, and is the reason your teeth feel slimy in the morning!
Bacteria cells exhibit one of several morphologies. The majority are either spherical (Cocci – Figure 1), rod-shaped (Coli – Figure 2), or spiral (Spirillia –
Figure 3).
Many bacteria remain as single cells, while others form colonies, the result of a single bacterium replicating. These colonies assume characteristic shapes
(chains, cluster of “grapes”), which can further classify the bacteria.
Figure 1: Cocci shape.
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Bacteria and Archaea
Bacteria do not undergo mitosis or meiosis like other cells. Each cell replicates its DNA and divides by fission or binary division. This results in identical copies of parent cells (clones). However, genetic variation may be introduced through recombination and mutation.
Small circular segments of DNA, called plasmids, enter the bacteria and integrate into its genetic code. These plasmids often code for genes that make bacteria resistant to antibiotics or viruses. Bacteriophages (often called just phages) are viruses that infect bacteria. Figure 2: Coli shape.
Figure 3: Spirillia shape.
Though we frequently consider bacteria to be harmful, many are not. In a healthy human body, there are 10-times more bacterial cells than human cells. Normally, these microbes are not invasive and do not cause disease (pathogenic). In fact, many benefit to the host by secreting vitamins, digesting food and performing other critical functions. When these bacteria are killed, as is the case with prolonged antibiotic treatment, serious health problems result. The bacteria that live on your skin can also cause disease if they enter the body through a wound or surgical incision (e.g., staph infections that occur after surgery result from staph on your skin getting inside of your body).
Bacteria also play an important role in our environment; binding nitrogen in plants, decomposing and recycling nutrients, synthesizing drugs and making food products (yogurt, sour cream). Bacteria have survived billions of years, in countless environments. Though scientists once thought they would eliminate diseasecausing bacteria, they continue to evolve and develop resistance to even the latest drugs (“anti”-”biotics”).
?
Did You Know...
The most prevalent chemical dissolved in the deep ocean waters near hydrothermal vents is hydrogen sulfide. The specialized bacteria that live near these vents use hydrogen sulfide as their energy source, instead of sunlight (chemoautotrophs).
The bacteria, in turn, sustain larger organisms in the surrounding area. The clams, mussels, tube worms, and other creatures at the vent have a symbiotic relationship with bacteria. They could not live without each other! The giant tube worms have no mouth or gut to digest food. Instead, the worm depends on the bacteria that have invaded its body to survive. There are around 285 billion bacteria per ounce of tissue within this tube worm. The plumes at the top of the worm's body are filled with blood, which contains hemoglobin that binds hydrogen sulfide and transports it to the bacteria that lives within the tissue. In return, the bacteria oxidize the hydrogen sulfide and convert carbon dioxide into carbon compounds that nourish the worm.
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Bacteria and Archaea
Pre-Lab Questions
1. Based on the scientific name Streptococcus agalactiae, what morphology would you expect these cells to have? 2. Name at least three animal structures that would be analogous to bacterial flagella.
3. Hypothesize how over-washing of hands can affect the population of “good” bacteria that resides on the human skin.
Experiment 1: Testing the Environment
Bacteria is found in almost all environments. Most are harmless, but there are those that cause disease. In this experiment, you will be test different environments for the presence of bacteria. You may be amazed to see how many different species are present on surfaces you come in contact with every day!
Materials
30 mL 8.25% Bleach Solution
*Hot Pad
4 Cotton Swabs (Sterile)
*Microwave or Boiling Water Bath
125 mL Nutrient Agar
*Refrigerator
60 cm Parafilm®
*Scissors (to cut the Parafilm®)
Permanent Marker
(4) 9 cm Petri Dishes
*You Must Provide
Notes About Working With Agar Plates…
•
Prepared agar dishes should be stored upside-down in the refrigerator until used. This will prevent condensation from disrupting the growing surface.
•
After inoculating, replace the cover on the dish, seal with Parafilm®, and store upside-down in a warm location (not to exceed 37.7 °C or 100 °F).
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Bacteria and Archaea
•
You should see growth within a few days. The plate will start to smell once microorganisms are growing. •
Before disposing plates, kill the microorganisms by pouring bleach solution onto the agar surfaces and let sit for 20 minutes.
Procedure
1. Loosen or remove the cap on the Nutrient agar bottle. Place in the microwave (if you do not have a microwave, place the bottle in a heat-safe bowl and pour boiling water around the bottle) and heat until the entire bottle of agar is liquefied. You will need to remove the bottle and swirl every 10 seconds to distribute the heat.
2. If you notice the liquid boiling over, STOP the microwave and let the bottle cool down before handling.
With a hot pad protecting your hands, remove the bottle from the microwave. Use caution when removing the bottle from the microwave as it will be HOT!
3. Gently swirl the bottle to mix the solution. Pour enough of the liquefied agar solution into the bottom half of 4 petri dishes so that it covers the entire bottom of the dish. Place the lids onto the dishes and CAREFULLY transport these plates into a refrigerator. Allow the plates to sit for 24 hours.
4. After 24 hours, remove the four agar plates from the refrigerator and allow them to sit at room temperature for at least one hour.
5. After the plates have warmed to room temperature, locate a surface to swab for bacteria. Things such as shoes, a table, teeth, bathroom doors, and shopping carts can be great sources. Try to avoid surfaces that are frequently cleaned with antibacterial detergents and soaps. On the bottom of the plate, write the name of the surface to be swabbed with your permanent marker.
6. Remove the lid from the agar plate. Unwrap one sterile cotton swab and smear the surface, being sure to roll the cotton swab to cover all sides.
7. Carefully streak the cotton swab onto the surface of the gelatinous medium (agar), being sure to start at the top and work down in a zigzag motion.
8. Place the lid onto the agar plate and seal it with a strip of Parafilm (hold one end of the Parafilm firmly against the side of the petri dish and stretch the other side to cover the entire perimeter). Place the plate upside down in a warm area to incubate.
9. Repeat this for two additional surface areas and agar plates.
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Bacteria and Archaea
10. For the last plate, label it “control.” Do not rub your sterile cotton swab on any surface, but rather take it straight from the package and streak it onto the plate. This will test for your accuracy in keeping the plates and swabs sterile while performing the experiment.
11. Let the plates incubate in a warm area for three days and then observe the growth. Answer the questions below.
12. After the experiment and after answering questions 1 - 3, set aside the plate with the most bacterial growth for the next experiment. Pour the bleach solution onto the surface of the remaining agar plates, allowing it to cover the entire surface. Let the plates sit untouched for 10 - 20 minutes. Then, seal plates with Parafilm® and dispose appropriately.
Post-Lab Questions
1. In the space provided below, draw your plates and indicate the number of different colonies and identify the colony shape.
2. Which plate grew the most bacterial species? Was this a surprise? Why or why not?
3. Was your control plate free of bacterial colonies? If not, how do you think the swabs were contaminated?
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Experiment 2: Measuring Antibiotic Resistance
In this experiment you will look at the ability of common antibiotics and other anti-bacterial agents to kill bacteria commonly found in your environment. You will use the bacterial lawn from the plate that produced the largest quantity of bacteria in the first lab.
Materials
1 Ampicillin Disc (marked)
1 Penicillin disc (marked)
10 mL 8.25% Bleach Solution
Permanent Marker
Forceps
Ruler
1 Kanamycin Disc (marked)
Parafilm®
1 Nutrient agar plate from the previous lab
Note: Be sure to wear gloves and use the forceps when handling the discs. Always pick them up if spilled.
Procedure
1. Using one of the plates covered with bacteria from the first lab, draw four even quadrants onto the bottom of the plate. Label them penicillin, ampicillin, kanamycin and control.
2. Wearing your gloves, goggles, and apron, open the plate and place the appropriate disks into the appropriate quadrants using forceps. Do not place anything in the control quadrant. DO NOT TOUCH THE
PLATE WITHOUT GLOVES.
3. Place the lid back onto the plate and cover it with Parafilm®. Allow it to incubate in a warm area for 3 days. 4. After 3 days, observe the zone of resistance from each disc (the region surrounding the disks where no bacteria grew). Using a ruler, measure these zones from the bottom of the agar plate.
5. After you are done with the experiment, pour enough bleach solution to cover the surface of the agar. Allow the plate to rest for 10 - 20 minutes. Seal the plate with Parafilm® and dispose appropriately.
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Post-Lab Questions
1. Did your bacterial lawn contain only one species of bacteria? If not, why do you think that is? Can you tell?
2. Which antibiotic was most effective in killing the bacterial lawn? Which was the least effective?
3. Each bacterial species shows different antibiotic susceptibility. What can you say about the bacteria that you grew?
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Lab 18
Protista
Pictured Above: Amoeba
Protista
Learning Objectives
•
Describe the diversity and complexity of organisms classified under Protista
•
Compare and contrast protist motility including pseudopodia, cilia, and flagella
•
Distinguish between protists that are autotrophic, heterotrophic, and mixotrophic
•
Distinguish between filamentous forming and colony associating protists
Introduction
Protists are some of the oldest and most diverse organisms on the planet. Most species which are grouped within the kingdom Protista share very few characteristics except for being eukaryotic. Some researchers refer to the kingdom Protista as a “catch-all” for organisms that do not fit into any other kingdom. Protists may be unicellular, multicellular or colonial; mobile or non-mobile; and possess mitochondria or chloroplasts.
Some protists are ever visible with the naked eye, although most require magnification.
Most protists reside in some type of water, whether it be the ocean, a pond, or within the human body. Their energy production can be either autotrophic or heterotrophic, with some switching between the two
(mixotrophic). All protists can reproduce asexually and some have the ability to reproduce sexually.
Protists can be defined as being unicellular, multicellular filamentous forming, or multicellular colony forming.
Chlamydomonas is a unicellular protist, more commonly known as green algae. There are over 600 species of this protist, but all contain chloroplasts which allow them to convert solar energy to chemical energy and provides them with their characteristic green color (Figure 1).
Figure 1: Chlamydomonas (green algae) are often found in stagnant water or damp soil.
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Protist Locomotion
Protists vary significantly in their motility. Some are entirely stationary, while others employ either flagella, cilia or pseudopodial movement. Flagella are common throughout the kingdom Protista, although they are also used by some prokaryotes. They are tail-like extensions that rotate or whip back and forth to produce locomotion. Flagellar movement may be due to a stimulus, such as a chemical gradient or food source. Depending on their location, flagella can either push or pull the protists through liquid medium. Some protists may have a single flagellum, while others have Figure 2: Giardia have two nuclei and four flagella. multiple flagella, like Giardia.
Ciliated protists use cilia, short, hair-like protrusions that enable the organism to move. The uniform motion of the cilia creates a “wave” that propels the protist. This wave also serves as a means for feeding by sweeping the food to them. One example of a ciliated protist is a paramecium. Other protists, such as amoebas use psuedopodial movement by extending a temporary protrusion called a pseudopod (Greek for “false foot”) to slowly crawl forward. Psuedopods are also used in feeding.
Slime Molds, Algae, and Protozoans Figure 3: Paramecium move using a coordi- The diversity of protists is vast, but this lab focuses on three prominent protists: slime molds, algae and protozoans. nated waving motion of their cilia.
Slime-Molds
Slime molds, often considered the link to the kingdom Fungi, share similarities with the kingdom Fungi such as appearance, nutrient acquisition, and the ability to form spores. In fact, some researchers refer to slime molds as “Fungus-Like Protists”. However, they differ in many key characteristics such as reproductive technique, life cycle and cellular make-up. Slime molds are phagocytic heterotrophs, meaning they surround and engulf particles with cytoplasm protrusions. They possess a complex reproductive cycle which includes mobile component (similar to that of animals), and an immobile component (similar to that of plants) when spore formation occurs.
Slime molds are either plasmodial or cellular. Plasmodial slime molds (Figure 4) are large, single-celled organisms with numerous nuclei. These nuclei are formed when many cells come together and fuse. Slime molds can sometimes be described as brightly colored cytoplasmic “blobs” which feed off of other microorganisms, such as bacteria. They can move via amoeboid movement, employing a method which mimics the pseudopodial movement of amoebas on a larger scale. They are quite useful in scientific studies because of their size.
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Cellular slime molds spend most of their life as individual single cells, but can come together and form a large multicellular organism. As the cells join, a slime covers the organism, allowing for uniform movement. Though the cells are attached, they do not fuse and remain distinct. They provide a useful tool to examine the formation of multicellular organisms and the ability of cells to recognize one another.
Algal Protists
With a multitude of colors, algae are often the most recognizable member of the kingdom Protista. Algae are photosynthetic autotrophs, meaning they possess chlorophyll and often other photosynthetic pigments. A select few are also heterotrophs, making them a mixotroph (an organism that has the ability to be photosynthetic as well as heterotrophic). There are three main types of algae: green algae, red algae and brown algae, each of which is distinct.
Figure 4: Plasmoidal slime molds typically span approximately three centimeters, but some have been documented to measure several meters in diameter!
Algae serve a number of important commercial functions. They produce over half of the oxygen in the
Earth’s atmosphere and are used for a number of commercial applications. For example, algae can be used to create ice cream, shaving cream, air fresheners, laxatives, dyes, pastry fillings, syrups and more).
Protozoans
Protozoans are typically unicellular organisms which require magnification to be seen. Like slime molds, they are phagocytic heterotrophs that surround and engulf small organisms. Since protozoans are motile, using psuedopodial movement, flagella or cilia, they are found in multiple environments. Though not all protozoans are harmful, as a group they are responsible for more disease and sickness throughout the world than any other group of organisms. Some of these diseases include Malaria (caused by the plasmodium protozoan), Giardia (caused by the giardia lamblia protozoan), African Sleeping Sickness (caused by the trypanosome brucei gambiense protozoan) and amoebic dysentery, also known as “Montezuma's Revenge” (caused by the entamoeba histolytica protozoan).
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Pre-Lab Questions
1. Hypothesize what type of environments make being autotrophic, heterotrophic or mixotrophic an advantage to a protist.
2. Many algae are photosynthetic like plants. Why then are they not classified as such?
3.Some protists are colony forming. Hypothesize why this may be an advantage or disadvantage.
Experiment 1: Viewing Preserved Species of Protists
In this lab you will look at preserved specimen slides of the Protist kingdom. While examining the slides, pay close attention to the individual characteristics of each organism (i.e. flagella, color, etc.).
Materials
Euglena Digital Slide Images
Paramecium Digital Slide Images
Procedure
1. Observe the detail of each organism (does it have flagella, what size is it, etc.). Look at Question 1. Draw each organism and label the major structures.
Post-Lab Questions
1. For each digital image (Figure 5 and 6), label the major structures you can see. What type of locomotion do you think this protist employs?
2. What advantages do you think a protist would gain by having a unicellular, filamentous, or colonial form?
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Figure 5: A digital slide picture of Euglena 1000X
Figure 6: A digital slide picture of Paramecium 1000x
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Experiment 2: Viewing Live Specimens
While observing the protists, you will notice that each member will exhibit different types of locomotion, speed, etc. You will observe a slowing reagent in part of the lab to get a better look at the fast moving protists.
Materials
Live protist specimen videos on the eScience Labs’ Student Portal
Procedure
1. Observe the movement of the protists with and without slowing agent. Notice the different types of locomotion employed by different types of protists.
Post-Lab Questions
1. In the space provided below, draw the various members of the protist kingdom that you were able to observe. Next to each drawing, describe the speed and type of locomotion you observed.
No slowing reagent
With slowing reagent
2. Did you notice a difference in the protists when the slowing agent was used? Which members were you able to see more clearly? Were there species that were still moving too fast to see clearly? Could you identify any species specifically? Do any of them look similar to specimens you observed in the first lab?
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Lab 19
Fungi
Fungi
Learning Objectives
•
Distinguish characteristics between the four major phyla of the fungi kingdom
•
Connect decomposers and saprophytes including hyphae and mycelium
•
Hypothesize why fungi have different and complex reproduction techniques including spores and dikaryon •
Describe the commercial uses of fungi
Introduction
Fungi are immobile, multicellular organisms (yeast is unicellular) that have some unique characteristics and play a profound role in ecology and human health. Fungi fall into four phylums:
•
Zygomycota (bread molds)
•
Ascomycota (sac- yeast and molds)
•
Basidiomycota (club mushrooms, rusts)
•
Deuteromycota (no identifiable sexual stage)
Fungi are decomposers that secrete enzymes to break down material into soluble organic compounds, which they can then absorb. They are saprophytes that digest dead and decaying material (e.g. ground, rotting wood, dung) and are critical in decomposing and recycling nutrients for the environment.
Multicellular fungi are primarily filamentous, with long, thin, somatic cells called hyphae. Unlike most plant and animal cells, hyphae can have more than one nucleus. Mycelium, a network of hyphae that lack distinct cells, are the structures that invade and digest material in soil, wood and other matter. Chitin is a light but strong substance found in the cell wall (in hard shells of insects and crustaceans) that adds rigidity and strength. Since the cytoplasms of hyphae are linked within the mycelium, a unique form of mitosis occurs in fungi. Instead of dividing nuclear material and the cytoplasm, mitosis occurs in the nucleus, which then divides (the nuclear envelope does not dissolve).
Reproduction
Spores are the basic reproductive structure of fungi. Fungi reproduce either asexually or sexually, depending on their environment. During the asexual phase, microscopic haploid spores are created by mitosis (the spores are identical). When they land in the right conditions, they germinate and grow new hyphae. There are five types of asexually produced spores:
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Fungi
1. Arthrospores, produced in fungi that have divided hyphae, are individual segment of the hyphae that break off into spores.
Figure 1
2. Chlamydospores, a segment of the mycelium, become thick-walled and breaks away as the spore. Figure 2
3. Sporangiospores are produced in a sac-like structure called the sporangium.
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Figure 3
Fungi
4. Conidiospores are not enclosed, but are produced in the conidiophores.
Figure 4
5. Blastospores are produced by a budding process (mitosis with unequal division of the cytoplasm).
Figure 5
Fungi form gametes through mitosis (unlike other multicelled organisms) and spend the majority of their life cycle in a haploid state (1n).
During sexual reproduction, hyphae of two different specimens, from the same species, meet and fuse together to form a dikaryotic cell (1n+1n). Once the nuclei have fused into a zygote, it goes through meiosis producing that mature into a new haploid organism.
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Fungi
Fungi produce three types of spores:
1. The fusion of two nuclei produces a thick-walled zygosporangium which manufactures “+” and “–“ haploid zygospores
Figure 6
2. Two fungi touch exchange nuclei that fuse together in a sac-like structure called the ascus. Meiosis then produces four nuclei which subsequently undergo mitosis, producing eight ascospores.
Figure 7
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Fungi
3. Mycelia fuse together to form a secondary structure with two nuclei. They maintain this dikaryotic state until fruiting occurs and the “gills” of the mushroom develop. Meiosis then produces four nuclei which migrate into the gills forming basidiospores.
Figure 8
Many fungi are harvested commercially for food, bread leavening, alcohol production, antibiotic production and other uses. Many symbiotic relationships have evolved between fungi and other plants and animals.
Fungi can produce chemicals that make food unpleasant, carcinogenic, or poisonous. They can cause athlete’s foot, ringworm and can kill plants (e.g. potatoes, elms and oak trees). Some are parasitic, feeding off living cells, absorbing their nutrients and killing their host.
Figure 9: Mushroom life cycle
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Definitions:
1. Chitin: A resilient and tough polysaccharide found in the cell wall of fungi, as well as the exoskeletons of insects and crustaceans
2. Dikaryon: Two cells that merge into one without fusing nuclei; a cell with two haploid nuclei
3. Hyphae: The branching, filamentous cells of fungi
4. Saphrophyte: An organism that feed on non-living organic matter
Pre-Lab Questions
1. Hypothesize why a fungus would use spores as a mechanism for reproduction.
2. How might the environment affect a fungus reproducing sexually or asexually?
3. What characteristics are more plant-like? Animal-like?
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Fungi
Experiment 1: Growing and Observing Zygomycota (Bread Mold)
Bread mold will generally ruin a day when you are looking forward to a sandwich, but will make it much easier to observe a live fungus. In this lab you will be cultivating the common bread mold. Any type of bread can be used, but fresh bakery bread will likely grow more colorful species since it does not contain the preservatives that many processed varieties do.
Materials
Permanent Marker
*2 Pieces of White Bread
Spray Bottle Lid and Empty Bottle
*Water
2 Resealable Bags
Rhizopus Digital Slide Images
* You Must Provide
Procedure
1. Use the permanent marker to label each of the two resealable bags as “Wet” or “Dry”.
2. Take one slice of bread and, spray it with water until moist. Avoid soaking the bread.
3. Place the moistened bread in the resealable bag labeled “Wet” and seal.
4. Then, place the second piece of bread in the resealable bag labeled “Dry” and seal.
5. Create a hypothesis regarding how the water will affect the bread over time. Be sure to address which piece of bread you think will host more growth, and why. Include your hypothesis in Post-Lab Question 1.
6. Incubate the bags for 3 - 7 days (depending on how fresh the bread is) in a dark, warm spot.
7. Examine the bread once mold starts growing.
8. Examine the digital slide pictures of Rhizopus (Figure 10) and identify the structures.
Figure 10: Rhizopus digital slide pictures; 100X (above left), 400X (above right).
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Fungi
Figure 10 (continued): 1000X
Post-Lab Questions
1. Include your hypothesis from Step 5 here.
2. What structures did you see in the bread mold?
3. Why was it important to moisten the bread before sealing it in the resealable bag?
4. What type of control (positive or negative) did the dry bread provide? Explain why.
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5. Is the bread mold reproducing sexually or asexually? How could you determine this?
6. What would you expect to find if you left the bread for six months?
7. Were multi-nucleated hyphae prevalent when looking at the Rhizopus slide? How do you know?
Experiment 2: Observing Ascomycota (Yeast)
Yeast is a commercially important member of the Fungi Kingdom. It leavens bread and produces beer and wine. Yeast is unique because it is unicellular and reproduces by mitosis and budding.
Materials
Yeast Digital Slide Images
Peziza Digital Slide Images
Procedure
1. View the digital slide image of yeast (Figure 11). Locate the budding cells and label what you see in the space below. Using the slide as a reference, draw a budding yeast cell and include important structures
(ascus, ascospore) on your picture.
2. View the digital slide image of Peziza (Figure 12). Identify the asocarp, asci, ascospores, and sterile hyphae.
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Figure 11: Yeast 1000X; field 1 (left) and field 2 (right)
Figure 12: Digital slide images of Peziza; 100X (above left), 400X (above right); 1000X (below)
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Post-Lab Questions
1. Compare yeast to the Peziza slide. Identify at least one difference, and one similarity.
2. How many ascospores are in each ascus of the yeast?
3. Are the ascospores of the Peziza inside or outside of the asci?
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Lab 20
Energy and Photosynthesis
Energy and Photosynthesis
Learning Objectives
•
Identify the four principle structures of a chloroplast: thylakoids, grana, lamella and stroma
•
Use photosynthesis to explain how the sun affects all organisms
•
Compare and contrast the light-dependent and light-independent reactions that occur during photosynthesis
Introduction
Photosynthesis is the process by which most plants, certain bacteria, and some algae harness light energy and manufacture food. In other words, it uses solar energy (light) to produce chemical energy
(carbohydrates). Photosynthesis consumes carbon dioxide (CO2, a greenhouse gas) and releases oxygen
(O2) (Figure 1). Nearly all life on Earth relies on this reaction in some format. Although there are many variants of photosynthesis found throughout nature, one of the most common products formed through photosynthesis is a sugar molecule called glucose (C6H12O6).
Figure 1: C6 Photosynthesis.
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Energy and Photosynthesis
All photosynthetic reactions are either light-dependent or light-independent. Both types occur in specialized organelles called chloroplasts in eukaryotes (plants and some algae species). The proteins involved in photosynthesis in photosynthetic bacteria are located in the cell membrane of these prokaryotes.
Chloroplasts, similar to that of mitochondria, are thought to have originated through an endosymbiotic event. That is, at some point in eukaryotic evolution, photosynthesizing bacteria (specifically cyanobacteria) were engulfed and became permanently associated with the eukaryote. In fact, chloroplasts maintain their own, unique circular double stranded DNA.
A chloroplast has four principal structures that play a role in photosynthesis (Figure 2):
Figure 2: Chlorophyll gives plants their green color. Wheat grass juice, common in health food stores, has a pronounced dark green color due to its high chlorophyll content.
1. Thylakoids: Small disk-shaped structures. These contain a pigment called chlorophyll which captures light energy.
2. Grana: Stacks of connected thylakoids.
3. Lamellas: Structures that link grana together.
4. Stroma: The fluid within the chloroplast, similar to cytoplasm. This fluid surrounds the grana and contains the liquids and molecules required for the Calvin Cycle. Chloroplast DNA and ribosomes are also located within the stroma.
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Energy and Photosynthesis
Plant Pigments
Plants have a number of different pigments which give plants their color and absorb light. Chlorophylls and carotenoids are two important classes of pigments. Chlorophylls are the greenish pigments and include several kinds.
Chlorophyll A is found in all photosynthesizing plants, algae and cyanobacteria. Chlorophyll B and chlorophyll C are less widespread and are only found in certain groups of organisms.
Carotenoids include the red, orange and yellow pigments. Carotenoids are sometimes referred to as accessory pigments because they pass the absorbed energy to chlorophyll when they absorb sunlight. Hundreds of known carotenoids are categorized into two major divisions: carotenes and xanthophylls. Beta carotene is the most abundant carotene in plants. This orange pigment gives carrots their color. Xanthophyll are yellow pigments and, similar to carotenes, are found in the leaves of most green plants.
Oxidation-Reduction Reactions (Review)
?
Did You Know...
Most of us have heard of the circadian cycles that humans engage in during sleep. But most people don’t know that circadian cycles also affect plant species to help them sense ambient light levels.
This cycle is regulated by oscillating biochemical processes which promote or inhibit the production of proteins involved in photosynthesis.
This regulation is critical to a plant’s ability to maintain its base anabolic actions which rely on solar energy.
Oxidation-reduction reactions, commonly referred to as redox reactions, are involved in photosynthesis. Redox reactions always include two separate reactions: an oxidation reaction and a reduction reaction. These reactions are so intricately connected that each oxidation and reduction reaction is often considered a halfreaction. Electrons are lost in oxidation reactions, and the same electrons are gained in the associated reduction reaction as the molecule transitions from the reactant to the product. When a molecule or atom undergoes oxidation, it is said to have been oxidized. Oxidizing agents are responsible for oxidation. Once the electrons are lost, the molecule is considered a cation. Cations are molecular ions which have a positive charge. A reduction reaction is one in which a molecule gains electrons as it transitions from the reactant to the product. Once the electrons are gained, the molecule is considered an anion. When a molecule or atom undergoes reduction, it is said to have been reduced. Reducing agents are responsible for reduction. Anions are molecular ions which have a negative charge.
Light-Dependent Reactions
Light-dependent reactions take place in chlorophyll A, chlorophyll B, and carotenoid pigments. Electrons act as the energy carrier throughout these reactions. Operating in tandem with other pigments, chlorophyll breaks the chemical bond of water and releases hydrogen and oxygen. These products, along with carbon dioxide undergo additional modifications in the stroma during the light-independent reaction stage (also known as the
Calvin-Benson Cycle). This ultimately results in the production of glucose, which can then be used for energy.
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Energy and Photosynthesis
Wavelength vs. Percent Absorbed of Different Plant Pigments
Figure 3: Wavelength vs. Percent Absorbed line graph. Violet: 400 nm, Blue: 475 nm, Green: 515 nm,
Yellow: 570 nm, Orange: 620 nm, Red: 675 nm.
Photosynthesis: White Light and Green Light
White light is a form of solar energy that contains all the wavelengths in the spectrum (i.e. “colors”: blue, green, yellow, etc.). Each pigment in a plant absorbs specific wavelengths of light, with most plants being unable to absorb green wavelengths. These are reflected back, which is why plants typically appear green. All of the other wavelengths (colors) are absorbed by the plant and initiate the formation of chemical energy. Figure 3 illustrates how each pigment absorbs specific wavelengths. Remember, color corresponds with the wavelength. Light-Independent Reactions (The Calvin-Benson Cycle):
After the light-dependent reaction, the light-independent reaction takes resultant products and converts them to carbohydrates and oxygen in a process known as the Calvin-Benson Cycle (Figure 4). The CalvinBenson Cycle is composed of multiple reactions that link carbon atoms to form simple sugar molecules such as glucose. This cycle begins when the enzyme RuBisCO (ribulose 1,5-bisphosphate carboxylase/ oxygenase) acquires carbon from the atmosphere in the form of carbon dioxide. Carbon fixation, reduction, and regeneration of ribulose then proceed to complete the Calvin-Benson Cycle.
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Energy and Photosynthesis
Figure 4: The Calvin-Benson Cycle.
Chromatography
Chromatography is a set of techniques that can physically separate components of a mixture (analyte) allowing for purification and analysis of these components. The physical and chemical properties of each analyte affect the rate of migration. For example, analytes with low solubility will migrate a shorter distance than analytes with higher solubility. These different rates, consequently, separate the analytes. In paper chromatography, the analytes are separated along a piece of chromatography paper.
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Energy and Photosynthesis
Paper chromatography is used to separate analytes, such as plant pigments, along a piece of chromatography paper. When the end of a piece of paper is dipped into water, the water molecules find new molecules to bond. The water molecules climb up the paper being replaced by new water molecules below. Solutes, which might be dissolved in the water, will also be carried along up the paper. The different solutes are carried along at different rates because they are not equally soluble. They are also differentially attracted to the fibers of the paper through the formation of intermolecular bonds, such as hydrogen bonds. Plant pigments can be separated by paper chromatography. After running the chromatogram, a retention factor (Rf factor) can be calculated for each analyte. The Rf is a ratio of the distance the analytes travel relative to the distance the eluting solvent travels.
The different pigments found in plants can be separated using paper chromatography. Beta carotene is able to travel relatively far up the paper because the pigment does not form any hydrogen bonds with the paper.
Xanthophyll, on the other hand, contains an oxygen group which forms hydrogen bonds with the paper.
Therefore, this pigment will be seen lower than beta carotene. Chlorophyll contains both oxygen and nitrogen, which bind even stronger to the paper and will travel even less than xanthophyll when separated with paper chromatography. You will be using pH indicators to detect carbon fixation, or the depletion of carbon dioxide, during the proceeding lab experiments. The concentration of carbon dioxide molecules decreases as carbon dioxide is reduced and oxygen is released. This causes the environmental pH to rise and the solution to become more basic. Pre-Lab Questions
1. Describe how the functional units for beta carotene, xanthophyll, chlorophyll A, and chlorophyll B are different. Be sure to identify the subunits that adhere to paper during chromatography.
2. Describe a technique for measuring photosynthetic rate.
3. Many deciduous trees have leaves which turn yellow in the fall. What do you suppose is happening in the leaves at the cellular and molecular level?
4. Chloroplasts and mitochondria are both are unusual in that they have double membranes and contain their own set of DNA. Can you think of any explanations for this observation?
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Energy and Photosynthesis
Experiment 1: Paper Chromatography
In this experiment, you will separate plant pigments using chromatography. You will also measure the rate of photosynthesis in isolated chloroplasts using the reduction of the dye 2,6-Dichloroindophenol (DPIP) as the measurement tool. The transfer of electrons during photosynthesis reduces DPIP, changing it from blue to clear. Materials
10 mL 4.5% Acetic Acid (Vinegar), C2H4O2
100 mL 0.5 M Sucrose Solution (cold!), C12H22O11
10 mL Acetone (Nail Polish Remover), C3H6O
3 Test Tubes (Glass)
30 cm Aluminum Foil
Test Tube Rack
(4) 100 mL Beakers
*Cutting Board
(1) 250 mL Beaker
*Kitchen Knife
20 cm Cheesecloth
*Light Source
3 mL 1% 2,6-Dichloroindophenol, DPIP
*Pencil
(1) 12 x 12 cm Chromatography Paper Piece
*Quarter
(1) 100 mL Graduated Cylinder
*Scissors
10 mL Mineral Oil
**Spinach Leaves (Fresh)
3 mL 0.1 M Phosphate Buffer, P
*Tape (masking or Scotch®)
8 Pipettes
*Water
(1) Resealable Plastic Bag
Rubber Band (Large; contain latex, handle with
*You Must Provide
gloves on if allergic)
*Keep these leaves if performing Experiment 2
Ruler
within a few days of Experiment 1.
Wooden Stir Stick
Procedure
Part 1: Paper Chromatography
1. Place the bottle containing 100 mL of sucrose solution in the refrigerator. Allow the solution to rest here for the remainder of Part 1; it will be required in Part 2.
2. Use the permanent market to label each 100 mL beaker as 1, 2, 3, or and 4.
3. Add 10 mL water to Beaker 1, 10 mL acetone to Beaker 2, 10 mL mineral oil to Beaker 3, and 10 mL acetic acid to Beaker 4. Cover the beakers with aluminum foil, and set them aside.
4. Use a pencil to draw a line approximately one cm from the bottom (across the bottom edge) of each piece
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Energy and Photosynthesis of chromatography paper. Then, carefully cut each piece in half so that you have four, pieces of identical size (each piece should have the pencil line going across the bottom edge of the paper).
5. Use a quarter to extract pigment from a fresh spinach leaf by placing the leaf in between the filter paper and the coin. Firmly rub the edge of the quarter back and forth over the pencil line drawn in Step 2. Use a fresh section of the leaf for each rubbing (Figure 5).
Figure 5
6. Place one piece of filter paper in Beaker 1, with the pigment-side towards the bottom of the beaker. Tape the filter paper to the beaker so that the bottom of the filter paper is submerged in the solvent, but the pigment line is not. DO NOT ALLOW THE PIGMENT TO TOUCH THE SOLVENT!
7. Monitor the set-up as the solvent travels up the paper. Remove the paper when the solvent is about 1 cm from the top of the paper. Immediately mark the solvent front and the location of each analyte band.
8. Measure the distance from the original pencil line to the solvent front, and the original pencil line to each of the pigments. Record the data in Table 1.
9. Calculate the Rf using the following equation:
10. Repeat Steps 6 – 9 for Beakers 2, 3, and 4. Be sure to use a new piece of filter paper for each beaker.
Rf = distance pigment migrated (mm) distance solvent migrated (mm)
Part 2: Chloroplast Isolation
11. Obtain a knife and cutting board. Carefully cut across a few spinach leaves to isolate chloroplasts from the leaves. 12. Place the leaves in a resealable bag. Measure and pour 100 mL of the cold sucrose solution (prepared in
Part 1: Step 1) into the resealable bag with the spinach leaves.
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13. Remove any air from the plastic bag and seal completely. Mash the solution for 2 minutes.
14. Lay a piece of cheesecloth over a 250 mL beaker, and push the cheesecloth into the beaker approximately four to five cm with your finger. Secure with a rubber band.
15. Slowly pour the spinach/sucrose solution into the cheesecloth. Allow the cheesecloth to filter the solution for several minutes; or, until all of the liquid has passed through the cheesecloth.
16. Once the liquid has drained, discard the collected solids. You may wish to squeeze the cheesecloth to extract any remaining liquid before throwing it away if liquid appears to be caught.
Part 3: Photosynthesis
17. Use the permanent marker to label three test tubes as 1, 2, and 3. Place the tubes in the test tube rack.
18. Add 3 mL water, 1 mL phosphate buffer, and 1 mL DPIP to each of the three test tubes.
19. Use the wooden stir stick to swirl the chloroplast solution
(prepared in Part 2). Then, add two drops of the solution to Tubes 2 and 3.
20. Immediately cover Tube 2 with aluminum foil.
21. Place Tube 1 and Tube 3 in a sunny location or under a strong light. If you place the tubes in front of a light, fill a beaker or clear glass with water and position in between the light source and the test tubes as shown in Figure 6.
Figure 6: A heat-sink for test tubes placed in front of a light source.
22. Monitor the test tubes for several hours, and record how long it takes each tube to turn from blue to clear. After the tubes exposed to light turn clear, remove the aluminum foil from Tube 2 and immediately note the color. Record your data in Table 2.
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Energy and Photosynthesis
Table 1: Part 1: Chromatography Data
Solvent
Distance from Original Line to Solvent Front
Number of Bands
Rf Factor
Acetic Acid
Acetone
Mineral Oil
Water
Table 2: Part 3 - Photosynthesis Data
Test Tube
Time Required to Change Color
1
2
3
Post-Lab Questions
1. What did the different colored bands signify in each solvent for Part 1? What pigments can you associate them with?
2. What is the osmolarity fluid used in Part 2? Why is this important? Why is it essential to keep it cool?
3. How could you modify this experiment to show the effects of different wavelengths of light on the photosynthetic rate?
4. Some plants (grasses) tend to contain a greater concentration of chlorophyll than others (pines). Can you develop a hypothesis to explain this? Would it be testable?
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Lab 21
Plant Circulation
Plant Circulation
Learning Objectives
•
Compare and contrast the structure and function of xylem and phloem in plant circulation
•
Hypothesize what factors affect transpiration rate
•
Explain the role water plays in transpiration
•
Compare and contrast transpiration with non-vascular water consumption
Introduction
Plants are multi-cellular, eukaryotes. They require energy and nutrients to engage in vital cell processes.
However, unlike animals which have a heart and blood to circulate critical elements, vascular plants must rely on their roots to uptake water and minerals from soil, and a specialized circulatory system to transport the nutrients to all other parts of the plant. This system is composed of xylem and phloem (Figure 1).
Xylem and Phloem
Xylem are hollow vessels produced from dead cells. They have additional support in their cell walls, which allows them to withstand the heavy weight imposed by the overall plant structure. Water and nutrients are transported from roots to leaves through the xylem conduits. Phloem are composed of living cells which form long, sieve-like tubes to transport the carbohydrate products of photosynthesis from the leaves to the rest of the plant. This process is called translocation. The vascular system allows plants to grow in diverse climates and to an extreme size. For example, Sequoia trees can live up to 2,200 years and grow to a height of 379 feet! That’s longer than a football field!
Figure 1: Root cross-section.
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Plant Circulation
Figure 2: Transpiration.
Transpiration
Water moves up the xylem, against the force of gravity. Water which reaches the surface of the leaves through the xylem may then evaporate and diffuse out of the stomata into the atmosphere in a process called transpiration. Stomata are small pore like objects which regulate the flow of water by opening to release water into the atmosphere through evaporation. There are millions of stomata in a single plant leaf.
When water evaporates, additional water and nutrients must be withdrawn from the ground to replace the water which has been lost. This process begins when water is absorbed by root cells and flows up the stem to the leaves and flowers, where it is allowed to escape (Figure 2). In this way, a continuous process of
“drinking” water from the soil and evaporation takes place.
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Plant Circulation
Figure 3: Evaporation process flow.
Transpiration Rate
A few key factors contribute to transpiration rate. These factors include humidity, light, temperature, and wind.
Humidity is inversely related to transpiration rate; in other words, increased humidity decreases the rate of transpiration. Light, temperature, and wind are all directly related to transpiration rate. For example, bright, windy, and high temperature conditions may all increase the transpiration rate.
Chemical and Physical Properties of Water
The chemical formula of water is recognized as H2O. Looking at the chemistry of the molecule, you can see that two hydrogen atoms are bound to one oxygen atom (Figure 4). However, the hydrogen atoms do not bind equidistant from each other.
Instead, two hydrogen atoms bond to the same side of the oxygen atom, creating molecular polarity. In other words, there is a positive charge on one side of the molecule, and a negative charge on the other. Since opposite charges attract, water molecules are attracted to other water molecules. In this form, or as a solid (ice), or gas (water vapor), water serves a great Figure 4: A molecule of water (red = oxygen atom; blue = hydrogen atoms). many purposes in the Earth’s atmosphere.
Due to the cohesive nature of water (polarity), as a water molecule leaves the plant, another takes its place.
The surface tension among the water molecules in the xylem, all the way down to the roots, ensures that as one molecule is lost to evaporation another replaces it in the roots. Think of sucking on a straw, as you suck liquid at the top, it is replaced by liquid at the bottom.
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Plant Circulation
Non-vascular plants (bryophytes) do not have a specialized circulatory system and are growth-restricted based on the availability of water. These low-growing plants, such as moss and liverwort species, usually thrive in dark, damp habitats where they can absorb water from their surface. Although they lack vascular tissues, true roots, leaves, and stems, they can transport water internally. Like sponges, they must imbibe the water needed for survival from their surrounding atmosphere through processes such as diffusion and capillary action. However, due to their limited terrestrial habitats, non-vascular plants have never dominated much of the Earth’s terrain.
Pre-Lab Questions
1. When did vascular plants evolve? How was this timeline uncovered?
2. How does a vascular system help a plant to grow bigger?
3. In 2 - 3 sentences, discuss how gas exchange affects transpiration.
4. Describe one way in which plants can adapt to water supply or environmental factors.
5. Why is water referred to as the “universal solvent”?
6. Why is a water droplet shaped like a sphere?
7. In essay form, discuss three adaptations plants have made to thrive in different environments.
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Plant Circulation
Experiment 1: Functionality of the Water Column
Materials
(2) 100 mL Beakers
flat on the bottom)
Permanent Marker
*2 Celery Stalks
Ranunculus root Digital Slide Images
*Cutting Board
15 - 20 Drops Red Dye
*Kitchen Knife
Ruler
*Water
Tilia Stem Digital Slide Images
*Bowl (Large enough to fit two pieces of celery
*You Must Provide
Procedure
1.
Examine the four Ranunculus root and Tilia digital slide images. In the space following the images, draw what you see. Label the xylem and phloem (refer to Figure 1 for help).
Xylem
Phloem
Figure 5: Ranunculus root (100X)
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Plant Circulation
Xylem
Phloem
Figure 6: Ranunculus root, 400X
Xylem
Phloem
Figure 7: Ranunculus root, 1000X
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Plant Circulation
Xylem
Phloem
Pith
Figure 8: Tilia (two year old stem; 40X).
Phloem
Secondary
Xylem
Primary
Xylem
Figure 9: Tilia (two year old stem; 100X).
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Plant Circulation
Phloem
Figure 10: Tilia (two year old stem; 400X).
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Plant Circulation
Include Your Drawings Here:
2. Fill a bowl with room temperature tap water.
3. Place the two celery stalks in the bowl. Use a knife to carefully cut ½ inch off the bottom of two celery stalks. This step should be performed with the celery underwater.
4. Leave one stalk submerged in the water in which it was cut, and let the other air dry for 15 minutes.
5. Use the permanent marker to label each of the 100 mL beakers as 1 or 2.
6. Fill Beaker 1 and Beaker 2 with 50 mL of water.
7. Place 8 - 10 drops of the red dye into each beaker.
8. After 15 minutes have passed, carefully, cut the tops off each stalk to create pieces which are approximately 10 cm. in length.
9. Place one stalk into each beaker with the freshly cut ends submerged in the water.
10. Observe the stalks for 12 hours, and answer the Post-Lab Questions.
Post-Lab Questions
1. How far did the dye travel in the celery which was submerged in water for 15 minutes? How far did it travel in the piece which was air-dried for 15 minutes?
2. In which celery stalk was the water column broken?
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Plant Circulation
Experiment 2: Water Movement in Flowers
Carnations colored green for St. Patrick’s Day or blue for the Fourth of July profit from transpiration to introduce color to the flowers. In this experiment, you will create your own colored carnation to gain a better understanding of how water travels through a plant.
Materials
(1) 100 mL Beaker
*Sharp Knife
8 - 10 Drops Red Dye
*Cutting Board
*You Must Provide
*1 White Carnation
Procedure
1. Fill a beaker with 50 mL water. Add 8 - 10 drops of the red dye. Under running water, cut the stem so that only one cm is left beneath the flower head. If this step is not performed under water, air bubbles can enter the xylem and prevent water from traveling to the flower.
2. Float the flower in the dye solution so that the fresh/exposed portion of the cut stem remains under water.
3. Set the beaker containing the flower in an illuminated place for 24 - 48 hours (depending on light intensity). Create a hypothesis to predict what will happen to the flower over the next two days. Record your hypothesis in the Post-Lab Questions section.
4. Observe the flower, and answer the Post-Lab Questions.
Post-Lab Questions
1. Record your hypothesis from Step 3 here. Be sure to address how the dye will affect the flower over time, and why.
2. In which part of the flower is the dye located? How did it get there?
3. What factors can influence the rate of transpiration in flowers?
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Lab 22
Plant Reproduction
Plant Reproduction
Learning Objectives
•
Compare and contrast vascular and non-vascular plant reproduction
•
Compare and contrast structure and function of bisexual and unisexual plants including gymnosperms and angiosperms
•
Identify plants as non-seed forming and seed forming
•
Explain alternation of generations during a plant’s life cycle
•
Apply double fertilization to angiosperms
Introduction
The plant kingdom encompasses many different types of plants. Embryophyta is a subkingdom of the plant kingdom which includes easily recognizable land plants such as mosses, ferns, flowering plants, and gymnosperms. Land plants have multicellular embryos which develop within the plant. They are further divided into vascular and non-vascular plants (or, bryophytes). Bryophytes encompass the oldest plant lineage, and are estimated to be approximately 475 million years old. Certain embryophytes form seeds to reproduce, while other types release spores. For example, bryophytes and some seedless vascular plants, such as fern, release spores. Vascular seed plants have adapted to a variety of habitats due in part to the robustness of the seed. Note that the common green algae is not an embryophyte. Unlike land plants which often (although not always) risk drying out, green algae is very hydrated as it absorbs water and minerals from its environment.
However, green algae can not be revived should it ever dry out, whereas many bryophtes are revivable if rehydrated within a sufficient time period.
Plant Life Cycle: Sexual Reproduction
The life cycle of all embryophytes, vascular and bryophytes, includes a diploid generation and a haploid generation. The diploid generation is called a sporophyte, and the haploid generation is called a gametophyte.
Diploid sporophytes produce hardy, haploid spores using meiosis. These spores become gametophytes by dividing and maturing. Gametophytes produce gametes by mitosis. Two gametes from different organisms come together and produce a zygote. The zygote develops into a sporophyte and the cycle continues. This cycle is the process for sexual reproduction in plants.
Alternation of Generations
The fluctuation between mitosis and meiosis is known as the alternation of generations. The time spent in each generation of the cycle varies by species within the plant kingdom. For example, the dominant generation in bryophytes is haploid. However, the dominant generation in most other plants is diploid.
There are similarities in how plants reproduce, but many differences are based on the size, form, and dispersion method of their gametophytes and sporophytes. For example, flowers, the sexual organs of angiosperms, vary greatly by size and shape, therefore requiring different methods of reproduction. Because plants
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Plant Reproduction are immobile, angiosperms (read more about angiosperms later in this lab) must utilize other, creative methods to seek out sexual partners.
Early plants including moss and ferns use water or wind to transport sperm for sexual reproduction. Further developed plants which possess formed vascular systems have adapted to other more complex methods of sexual reproduction. For example, some flowers express brightly-colored attract insects to cross-pollinate. Other flowers express caffeine molecules in their nectar. This caffeine is thought to improve memory in honeybees, which improves the flower’s likelihood Figure 1: Dandelion releasing seeds. of repeat pollinations.
Unisexual vs. Bisexual Reproduction
Plant reproduction can be unisexual or bisexual. Bisexual reproduction occurs when a plant incorporates structures which are both male and female. Unisexual reproduction occurs when a plant incorporates structures which are either male or female.
Mosses and liverworts are unisexual plants that do not form seeds to reproduce
(Figure 2). This class of plants has developed both male and female reproductive organs. The female organ is the archegonium where the archegoniophore (egg or female gametophyte) resides. The male organ is the antheridium where the antheridiophore (sperm or male gametophyte) resides. The antheridiophore depends on water for movement (dew, rain) and male and female plants must be in close proximity to each other for successful fertilization.
Gymnosperms
Gymnosperms (e.g. pine trees) are bisexual plants that produce naked seeds. Gymnosperms are called the naked seed plants
Figure 2: Moss reproduction cycle. because they are not encased by the mature ovary (fruit) as other seed-forming plants are. Although each plant contains both the male and female reproductive structures, they are in physically different locations. Typically the female reproductive structures
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Plant Reproduction
(seed cones) are near the top of the tree while the male reproductive structures (pollen cones) are near the bottom of the tree. These plants are called “wind pollinated” because they rely on the wind to blow pollen grains (male gametes) to the ovule that contains the female gametes. Pollination occurs when the pollen grains come in contact with the ovule. The pollen grains germinate and pollen tubes will grow towards the egg. Once the tube reaches the egg, sperm are released from the grain and fertilize the egg. The zygote
(seed) will then mature and fall from the seed (Figure 2).
Angiosperms
Angiosperms, or flowering plants, are vascular, bisexual plants. Both the male and female reproductive organs are found in a single flower. While these plants can selffertilize, cross-fertilization (pollen from a different plant) is important to maintain genetic diversity. The carpel is a specialized organ, unique to angiosperms, that encloses the ovule and eventually matures into fruit following fertilization (Figure 3). Angiosperms are called covered seed plants because the developed seed (zygote/embryo) is within the fruit.
Angiosperms have additional distinct reproductive organs called flowers. The gametophyte is reduced to the female embryo sac, with as few as eight cells and the male gametophyte developed from the pollen grains.
Pollen grains, the male gametophyte, developed to protect the sperm during the transfer from male to female
Figure 3: Flowering plant reproductive organs. parts. Double-fertilization is also unique to these plants.
Like the gymnosperms, a pollen grain that reaches the carpel will germinate. Pollen tubes grow towards the ovary and two sperm are released. One sperm nuclei fuses with the egg to form the zygote (2n). The other nuclei unites with two haploid polar cells (in the ovary) to form a triploid (3n) cell that develops into the endosperm, a nutrient-rich tissue surrounding the seed (fruit).
Pre-Lab Questions:
1. Identify one reproductive difference between sporophytes and gametophytes.
2. How do gymnosperms vary from other, seed-forming plants?
3. How do abiotic factors such as wind or water influence pollination behavior?
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Plant Reproduction
Experiment 1: Observation of Archegonium and Antheridium
Mosses are the most common group of unisex, non-vascular plants, and are common in moist environments.
Long sections of the moss Mnium containing the gametangia are useful to gain a better understanding of the moss life cycle. The antheridium is a haploid structure involved in the production and storage of sperm cells.
Alternatively, the archegonium is a structure involved in the production and storage of ova.
Materials
Mnium moss Archegonium Digital Slide Images
Mnium moss Antheridium Digital Slide Images
Procedure
1. Examine the digital slide pictures of the archegonium (Figure 4, 5, and 6). Look for the archegonium, a structure with a long neck and rounded base. Also, search for a single-celled egg within the archegonium.
Label those structures.
2. Examine the digital slide picture of the antheridium (Figures 7, 8, 9, and 10). Sperm-forming tissue will be visible within the antheridia. Label significant structures.
Egg
Archegonium
Figure 4: Mnium archegonium (100X).
Post
-Lab Questions
1. Are the spores produced by the moss sporophyte formed by meiosis or mitosis? Are they haploid or dip314
loid?
Plant Reproduction
Egg
Archegonia
Figure 5: Mnium archegonium (400X).
Egg
Archegonia
Figure 6: Mnium archegonium (400X).
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Plant Reproduction
Gametophyte
Tissue
Antheridia
Figure 7: Mnium antheridia (40X).
Antheridia
Figure 8: Mnium antheridia (100X).
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Plant Reproduction
Sterile Jacket
Antheridia
Spermatogenous Tissue
Figure 9: Mnium antheridia (400X).
Sterile Jacket
Antheridia
Sperm Forming Tissue
Figure 10: Mnium antheridia (1000X).
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Plant Reproduction
Experiment 2: Observation of a Flower
In this experiment you will observe slide images of flower reproductive parts and compare them to an actual specimen, Materials
Lilium Ovary Digital Slide Images
Lilium Anther Digital Slide Images
*You Must Provide
*Fresh Flower (Lily is recommended)
Procedure
1. Study the fresh flower. Locate and identify the important parts of the flower’s reproductive system. Sketch the flower in the space below; be sure to label the reproductive components.
2. Examine the digital slide images of the anther and ovary. Pay special attention to the labeling and review the annotations.
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Plant Reproduction
Anther sac with pollen
Figure 11: Lilium anther (40X).
Pollen Grains
Pollen Sac
Figure 12: Lilium anther (100X). Pollen grains (microspores) are clearly visible within the pollen sac (microsporangium).
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Plant Reproduction
Pollen Grains
Pollen Sac
Figure 13: Lilium anther (400X).
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Figure 14: Lilium anther (1000X).
Plant Reproduction
Pollen Grains
Ovum
Ovule
Ovary Wall
Pollen Sac
Figure 15: Lilium ovary (40X).
Embryo
Sac
Ovules
Ovary Wall
Figure 16: Lilium ovary (100X).
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Plant Reproduction
Ovule
Embryo
Sac
Figure 17: Lilium ovary 400X.
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Plant Reproduction
Post-Lab Questions
1. What are some unique characteristics of angiosperms?
2. How many carpels are in the lily? How many stamen?
3. Describe the male gametophyte of a seed plant.
4. Describe the female gametophyte of a seed plant.
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Lab 23
Invertebrates and Vertebrates
Invertebrates and Vertebrates
Learning Objectives
•
Compare and contrast the major structures and functions of invertebrates and vertebrates
•
Identify phyla as invertebrates or vertebrates: Porifera, Cnidaria, Platyhelminthes, Annelida, Arthropoda, Mollusca, Echinodermata, Chordata and Vertebrata
•
Identify structural differences and similarities between major classes of vertebrates
•
Classify organisms as displaying radial, bilateral symmetry or asymmetry
Introduction
The kingdom Animalia comprises millions of species, ranging from the snail to the hippopotamus, the ant to the elephant, the centipede to the human. Though there are notable differences in body shape and function, almost all species are motile, multicellular eukaryotes. Unlike plants, animals cannot produce energy from sunlight and, as heterotrophs, they acquire energy by consuming organic material (other plants and animals).
Symmetry
Symmetry is a useful characteristic for animal classification and represents the balanced division of an animal’s form. Radial symmetry, as is seen in starfish, is a division originating in the center and protruding outwards that produces even and balanced sections. This is similar to the divisions that are made when a pie is cut into many even pieces. Bilateral symmetry, as is seen in beetles, occurs when animal structure can be divided into two mirror images by a center line that runs through the entire object (Figure 1). No equal divisions can be detected in asymmetry, as is seen in a sponge.
Invertebrates
Animals are classified as invertebrates or vertebrates based on their internal structure. Invertebrates are organisms that lack a backbone. They make up over 98% of all animal species. Examples include the jellyfish, insects, or worms. Vertebrates pos- Figure 1: The rhino beetle exhibits bilateral sess an endoskeleton (an internal skeletal structure) and a spi- symmetry along the central axis. nal column. A backbone or spinal column is a significant adaptive advantage that enables vertebrates to occupy different ecological niches (roles). Examples include humans and dogs.
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Invertebrates and Vertebrates
Major Invertebrate Phyla
There are over 35 phyla in the Animal Kingdom. The vast majority of organisms fall within the following eight phyla, seven of which are invertebrates.
1. Phylum Porifera consists of sponges, considered to be the oldest of the animal phyla (Figure 2). As stationary filters, sponges play an important role in aquatic systems, feeding on particles and bacteria in the water. Porifera literally means pore bearing which accurately describes the exterior surface of the sponges.
Water enters the sponge through small pores and exits via larger openings. Nutrients are filtered out during the unidirectional flow of water through the sponge. Their asymmetric body is a loose assembly of cells (no tissues) that is supported by a matrix of collagen fibers and spicules. Sponges reproduce both sexually and asexually. 2. The phylum Cnidaria includes jellyfish, corals, sea anemones, and hydras (Figure 3). They are thought to be the first animals to develop nerves and muscles and typically alternate between two body forms: the free-swimming medusa and the stationary polyp. Both body types consist of three layers of tissue surrounded by tentacles with stinging cells containing tiny, toxic harpoons that can be used in either defense or offense.
In fact, the name “cnidarian” is derived from the Greek word for stinging nettles. They exhibit radial symmetry with a hollow body cavity to digest food. Like sponges, cnidaria can reproduce both sexually and asexually.
Figure 2: Porifera
Figure 3:Cnidaria
3. The phylum Platyhelminthes (flatworms) includes freshwater planaria, colorful marine polyclads, parasitic tapeworms and flukes. They are some of the simplest bilaterally symmetrical organisms with a defined head and tail, and a centralized nervous system containing a brain and nerves.
They lack both a body cavity and circulatory system, but do have a tubular mouth, an excretory system and a highly branched digestive system. Clusters of light-sensitive cells make up their eyespots. Recent molecular studies using DNA sequences have aided in the classification of many of these organisms which may eventually lead to a reorganization of this phylum. They are hermaphroditic, capable of both sexual and asexual reproduction.
4. The phylum Annelida is represented by marine worms (polychaetes), earthworms and leeches (Figure 4). They are bilaterally symmetric with segmented body cavities. These cavities are often represented as a tube within a tube design. Each segment has tiny hairs or bristles called setae which help the organism to move. Segmentation was an important development
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Figure 4: Annelida
Invertebrates and Vertebrates that provides distinct regions to specialize in different tasks. A one-way digestive tract, closedcirculatory system, and central nervous system also differentiate this invertebrate. Annelids play a significant ecological role by reworking soil and sediments. Annelids primarily reproduce by sexual means with some members capable of both sexual and asexual reproduction.
5. The phylum Arthropoda, the most diverse and numerous of the Ani-
mal Kingdom, includes insects, crustacean, spiders, millipedes, and centipedes (Figure 5). All arthropods have segmented bodies and are covered in a hard, flexible exoskeleton. The muscles from their jointed appendages attach to the inside of this protective cover. Many species, such as dragonflies which start as larvae and develop into winged adults, exhibit multiple life cycles. Their open body cavity contains tissues, organs and a complete digestive tract. The vast majority Figure 5: Arthropoda of members of arthropoda reproduce sexually.
6. The phylum Mollusca includes clams, snails, slugs and the octopus (Figure 6). Though there is a great deal of diversity within this phylum, all mollusks have soft bodies, many of which are covered by a hard calcified shell. The shell is produced through secretions from a layer of tissue called the mantle (also known as the pallium). A muscular foot provides locomotion and grasping. A coarse, file-like organ (the radula) allows most mollusks to drill into their prey or snag fish. Many hunt by propelling water through a siphon either for locomotion or to capture food. A mantle cavity houses gills and one-way digestive system. Ongoing molecular studies may lead to a reorganization of this phylum as more information becomes available. Most mollusks have separate sexes and reproduce sexually, although some mollusk speFigure 6: Mollusca cies are hermaphrodites.
7. The phylum Echinodermata includes sea stars, sea lilies, sea urchins, sea cucumbers, and over 6,000 other salt water species (Figure 7). Instead of bilateral or radial symmetry, most echinoderms exhibit a five part morphological symmetry. Their hard, flexible bodies are composed of small calcium plates that are often spiny and covered by a thin skin.
In fact, the name “Echinodermata” means spiny skin. Internal to the spiny skin, there is a complete digestive system and a special fluid-filled system that operates tube feet (which sometimes grow back if lost) which allow them to move, feed, and respire. Most organisms in this phylum reproduce sexually with a few specific species known to reproduce asexually. Figure 7: Echinodermata
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Invertebrates and Vertebrates
8. The phylum Chordata includes organisms that are invertebrates and organisms that are vertebrates unlike the above seven phyla which include exclusively invertebrates. Although it may seem unusual to have a phylum containing both invertebrates and vertebrates, there are four very specific embryonic characteristics or traits that allow for this classification. That is, all chordates possess each of these traits at some point during their embryonic development. First, all chordates develop a notochord. This rod-like, supportive structure extends the length of the body. Second, a dorsal nerve cord running parallel with the notochord is established. The last two traits are the development of both a muscular tail that extends beyond the anus and gill slits.
Chordates have bilateral symmetry, segmented muscles, a protective layer (feathers, scales, hair, fur, etc.,) and gill clefts (the structures located behind the mouth and in front of the esophagus).
The invertebrate chordates are the tunicates and the lancelets, both are types of marine organisms. Tunicates, more commonly known as sea squirts, look like sponges but have a notochord with a dorsal nerve cord gill slits and a tail. Lancelets are small fish-like creatures that possess the embryonic structures allowing for the classification of chordate. There are over 20 different species of lancelets.
Keep in mind that, as with many classifications, some vertebrate species might be included within the subphylum chordate in some resources, but not in other resources. For example, hagfish do not have true vertebrae but they do have a cranium. Based on the lack of true vertebrae, hagfish do not technically belong in this subphylum, but they often are classified within this group. To circumvent this issue, the term Craniata is frequently used. Craniata include animals within Chordata that have a skull.
Vertebrates
Vertebrates possess an internal skeletal structure called an endoskeleton and a spinal column. This is a key differentiating feature between vertebrates and invertebrates, which do not possess a hardened endoskeleton.
An endoskeleton is a significant adaptive advantage that enables vertebrates to occupy different ecological niches (roles) and engage in a variety of unique animal behaviors. Examples of the over 60,000 vertebrate species include humans, dogs, and the llama shown in Figure 8.
Although vertebrates and invertebrates are very different for many reasons, there are four very specific embryonic traits that all chordates possess at some point during their development. These traits include: the development of a notochord, a dorsal nerve cord, gill slits, and a muscular tail that extends beyond the anus.
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Figure 8: Llamas are historical companions to mountain trekking humans. They are often referred to as pack animals due to their ability to distribute weight over their vertebrae and back.
Invertebrates and Vertebrates
Fish and Tetrapods
The two main groups of the subphylum vertebrata include fishes and tetrapods (amphibians, reptiles, birds, and mammals). Fish are aquatic and lay soft eggs. They are cold blooded and use gills to breathe. As the name suggests, tetrapods include all animals with a four-legged body plan such as mammals and amphibians. Amphibians, such as frogs and salamanders, are found in both water and on land and lay soft eggs. Amphibians are cold blooded and breathe using gills, lungs and through their skin.
Amniotes, Reptiles, and Birds
The development of amniote eggs provided animals (amniotes) with the capability to move away from water.
Amniote eggs possess internal membranes that keep embryos moist. The two major divisions within amniotes are the reptiles (non-bird reptiles and birds) and mammals. Reptiles live almost entirely on land and lay somewhat hard shelled eggs. They are cold blooded and breathe through lungs. Birds are found on land
(and in the air) and lay hard shelled eggs. Birds are warm-blooded and breathe through lungs.
Mammals
Mammals, including humans (Homo sapiens), are normally found on land and most give live birth. They are warm-blooded and breathe through lungs. Mammals are also characterized by the presence of hair on their bodies and their ability to produce milk in modified sweat glands
(mammary glands) for their young. There are roughly 5,000 species of mammals.
There are three mammalian groups or lineages. Monotremes are mammals that lay eggs and appear more similar to reptiles with their somewhat leathery shell. Monotremes produce milk which is excreted through openings found on the mother’s skin due to the absence of nipples. The duck-billed platypus and two types of spiny anteaters are the only three monotremes species.
Marsupials are pouched mammals and include kangaroos, koalas and opossums. Marsupials give live birth to developmentally premature (or, altricial) young. The young make their way to the mother’s ventral pouch. They continue developing while being nourished by milk supplied by the mother through nipples located in the pouch.
Placental mammals develop an organ, called the placenta, during pregnancy which acts to transfer nutrients and gases between the mother and the embryo. Placental mammals include primates, all marine mammals and rodents. In fact, over half of the Figure 9: Newborn joeys are typically apover 4,000 placental mammals are rodents. proximately the size of a jelly bean.
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Homo sapien
Arguably the most dominant vertebrate is Homo sapien (humans). Humans are thought to be some of the longest living mammals, though other species, such as the elephant and whale are also long-lived. Though there are obvious differences between human beings and other placental mammals, there are also many similarities. Major differences include skeletal changes that have allowed for humans to stand and walk upright on two legs (bipedalism). Being bipedal also allowed for hands to become adapted to more specialized and novel roles. Human backbones are slightly S-shaped, which helps keep the body aligned over the hips. The human skull also differs so that it sits on top of the backbone with the spinal cord exiting the skull inferiorly.
Figure 10: Human spines are naturally S-shaped. However, excess curvature may result in scoliosis.
Pre-Lab Questions:
1. Identify the four major characteristics of chordates.
2. List ways in which invertebrates compensate for not having a spine.
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Invertebrates and Vertebrates
Experiment 1: Symmetry in Common Objects
Symmetry can be seen in everyday objects similar to that of the symmetry seen in organisms.
Materials
Images of Common Lab Equipment
Procedure
1. Review the objects listed below (many of these can also be found in your lab kit).
2. Indicate the type of symmetry each item displays next to the image in the Post-Lab Questions section.
Hint: It is helpful to draw lines of symmetry over the objects and observe objects from different angles.
3. For each item, explain why you chose the type of symmetry you did.
a. Petri Dish
d. Wash Bottle
b. Funnel
c. Test Tube Rack
e. Graduated Cylinder
f. Tongs
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Invertebrates and Vertebrates
Post-Lab Questions
1. Identify the type of symmetry displayed for each item. Then, indicate how you came to your conclusion:
a. Petri Dish:
b. Funnel:
c. Test Tube Rack:
d. Wash Bottle with Curved Straw:
e. Graduated Cylinder:
f.
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Tongs:
Invertebrates and Vertebrates
Experiment 2: Creating a Phylum Key
In this experiment you will take a closer look at the characteristics that describe the eight major phyla.
Materials
Table 1
Table 2
Procedure
1. Table 1 contains all of the main features of the eight major phyla discussed in the Introduction. Organize and identify which characteristics belong to which phyla in the key. The number of rows in the key represents the number of characteristics that fall into each phylum.
2. Record your results in Table 2.
Table 1: Phylum Characteristic Table
1
2
3
4
5
A
Bilateral phylum with segmentation
Most have a calcium containing shell
Five part symmetry
Specialized cell, but no tissues
Mantle of tissue covering the body
B
Hollow body cavity for food
Three tissue layers, no body cavity
Setae used for movement Jaws and skulls part of evolution
Complete digestive tract
C
Parasites
Water flows through canals of body
The first to have jointed legs
The first phylum to fly
Some have stinging cells
D
More complex because of more DNA
Body design is a tube within a tube
Tube feet
First muscle and nerves Internal skeleton
E
All live in the sea Simple animals with bilateral symmetry
Entrance and exit the same in the digestive tract
All have vertebral column
Champions of variations in appendages F
Has the most species Have spines covered with a thin skin
Some stationary polyps Some are mobile medusa Some propel using their siphon G
Muscular foot used to move
Stationary animal
Tubular mouth at the mid-body
Humans
Hard but flexible bodies with small plates
H
Spicules are the skeleton
Radula used to feed
No symmetry
Exoskeleton
Their burrowing has affected the global climate
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Invertebrates and Vertebrates
Table 2: Phylum Taxonomic Key
Porifera
Cnidaria
Platyhelminthes
Annelida
Arthropoda
Mollusca
Echinodermata
Chordata
Post-Lab Questions
1. Were any of the features used in more than one Phyla? If so, give an example of an organism from each phyla that shares the feature.
2. List three features from Table 1 that describe a Monarch butterfly (Danaus plexippus)? Using the features you listed determine what phyla it is classified in.
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Invertebrates and Vertebrates
Experiment 3: Taxonomy
Materials
(6) Organism Images
Procedure
1. Identify which phylum each of the following organisms belongs in. Next to each, list the criteria used in your determination.
a.
b.
c.
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Invertebrates and Vertebrates
d.
e.
f.
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Invertebrates and Vertebrates
Experiment 4: Owl Pellet Dissection
Birds of prey often consume their prey items whole, rather than selecting for the digestible components.
However, proteolytic enzymes present within the bird’s stomach are not capable of digesting all of the different structures found in an organism, and often can’t break down items such as hair, bone, teeth, bones, or exoskeletons. As a result, owls (like many other predatory bird species) produce and regurgitate pellets which contain many of these indigestible items approximately 18 - 20 hours prior to consumption. Very small bones or bone fragments occasionally pass through the pyloric sphincter and proceed through the digestive system, but pellets often contain complete animal skeletons.
In this experiment, you will probe a barn owl (Tyto alba) pellet to learn more about the skeletal system’s general physical characteristics. Note that some of the bones which you recover from the pellet will appear homologous to a human skeleton (e.g., the ribs and vertebra), while others will vary significantly (e.g., the skull). Materials
Construction Paper (Black)
*Paper Towel
Forceps
*Soap
Hand Lens
*Water
Owl Pellet
Ruler
*You Must Provide
2 Toothpicks
Caution: The owl pellets have been heat-treated but may still harbor microbes. Wear protective glasses, gloves, and apron when performing this experiment. Thoroughly wash your hands and all work surfaces with warm water and soap after complete.
Procedure
1. With your gloves on, unwrap the aluminum foil from the owl pellet and set the pellet atop the black construction paper. This will make it easier to identify the bones in the pellet as you dissect.
2. Measure the dimensions of your pellet. Record your data in Table 3.
3. Review the physical characteristics of the pellet such as texture, color, scent, etc. Be sure to use the wafting technique to detect the scent of the pellet (see the Appendix for a description of this technique).
Record your observations in Table 3.
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Invertebrates and Vertebrates
4. Carefully use the toothpicks and forceps to probe the pellet. If your pellet feels very firm, submerge it in a beaker filled with water for 1 - 3 minutes to soften it.
5. Isolate any bones you come across.
6. Continue to probe the pellet for the bones, removing fur and debris from the bones. Note that the bones will be fragile. Be careful not to break any of the bones during the dissection.
7. Use Figure 12 to identify the bones found in the pellet. Record the bones you identify in Table 3.
8. Try to recreate the skeleton of one animal found in the pellet by organizing the bones in the arrangement of the skeletal system. Pay particular attention to where the joints might have been located.
Note: Each pellet is likely to contain bones from several different animal species. Be sure to refer to
Figure 8 to complete this step.
9. Dispose of the pellet in a trash receptacle in a location which is safe from children or animals. Then, wash your work station with warm water and soap.
Table 3: Owl Pellet Data
Pellet Characteristics
Pellet Length (cm):
Pellet Width (cm):
Physical Observations:
Bone
Skull
Jaw
Scapula
Rib
Vertebrae
Hindlimb
Forelimb
Pelvic bone
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Animal Source
Number of Bones
Invertebrates and Vertebrates
Figure 12: Owl pellet bone identification chart.
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Post-Lab Questions
1. What did you find in your owl pellet?
2. What have you learned about the ecosystem in which the owl lives?
3. What can you infer about the nature of the community in which the owl lives?
4. How can scientists use owl pellets to study small mammals in a specific ecosystem?
5. Other birds of prey produce pellets as well, and the contents are dictated by where the bird lives. What would you expect to find in the pellet from a shorebird, such as a gull?
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The following lab exercise is intended to be an add-on for the Invertebrates Lab.
You will NOT have the supplies to perform the next experiment unless your teacher has specifically requested that dissection materials be included within your kit.
Experiment 5: Invertebrate Dissection
The starfish is not a fish at all, rather an invertebrate that possesses no internal skeleton. Members of the phylum Echinodermata, starfish are unique in that they are deuterostomes (instead of protostomes like earthworms, grasshoppers, clams, etc.). Deuterostomes exhibit incomplete segmentation, and a brain and spinal cord above the gut, among of other differences with protostomes. Starfish have no front or back, and can move in any direction without turning.
Note: When performing a dissection, remember these important safety notes:
•
Dissect with the scalpel or scissor blade cutting away from you (and your lab partner).
•
If you develop an allergic reaction to the preserving fluid, contact your healthcare provider. Also, inform your instructor and inform him/her of your situation.
•
Contact your local waste management company for instructions for the proper disposal of your specimen. •
Wash your hands, dissection tools, and all work surfaces with soap and water when finished with the dissection.
Materials
Dissecting Pins
Hand Lens
Dissecting Tools
Preserved Starfish
Dissecting Tray
Procedure
1. Examine the external anatomy of the starfish. The side in which the mouth is located is called the oral surface (ventral side). The opposite side is called the aboral surface (dorsal side). See Figure 13 for reference.
2. Run your gloved finger over the surface and note the texture. Use the magnifying glass to examine the spiny skin in detail.
3. Along with the stiff spines, you may also see small, hair-like gills used by the starfish to take in oxygen.
4. Pedicellaria are tiny pincers that look like pliers that are used to grip small objects.
5. Place the starfish ventral side up and note the tube feet that run down the arms, or rays, of the starfish.
They will be located on either side of the groove that runs from the tip of each ray to the center.
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Figure 13: The aboral (left) and oral (right) surfaces of the starfish.
6. Find the mouth in the center of the starfish.
7. On the oral surface of each arm are open ambulacral grooves extending from the mouth to the tip of each arm. Locate the abulacral groove running from the center down each ray.
8. Using the magnifying lens, examine the tube feet with protruding suckers on either side of the abulacral groove. 9. Flip the starfish so the dorsal side is facing up.
10. Note the eyespots at the tip of each arm, which allows the starfish to sense and respond to light. Eyespots are pigmented regions near the tip of the arms, and are covered by a transparent cuticle. To see the eyespots, spread the tube feet at the tip of the ray and examine it closely with the magnifying lens.
11. The flat central disk is at the center, and tiny, hollow, finger-like gills cover the body of the starfish.
12. The opening of the water-vascular system is called the madreporite. It is a large, button-like structure on the central disk.
13. The anus is in the center of the disk.
14. Using a scalpel, cut one inch from the tip of one of the rays. Study the cross section of the stump.
15. Note the ossicles (part of the endoskeleton) on the dorsal surface, the largest of which called the ambu-
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Invertebrates and Vertebrates lacral ossicles, which support the ambulacral groove and provide attachment for the tube feet.
16. Remove the skin from the top of this ray using dissecting scissors or scalpel. Do the same for another ray, and also cut a circular lap of skin from the central disk being careful to keep it as shallow of an incision as possible. 17. Note the feathery-looking digestive glands called the pyloric caeca. These glands make enzymes that help digest food in the stomach, located under the central disk. The thin sac lying just above the stomach is the intestine. From there, the rectal pouches store small amounts of wastes before leaving through the anus on the dorsal side of the starfish.
18. Remove the pyloric caeca from one ray, and observe the gonads underneath.
19. Remove the gonads to visualize the water vascular system. This is an internal water pressure system.
Water enters the system through the madreporite, passes through a series of canals until it reaches the tube feet. When the ampulla contracts, water is forced into the tube foot, extending it and allowing it to grab on to a surface.
20. Running the length of each ray is a lateral canal, to which tube feet are attached.
21. In the central disk, the five lateral canals connect to the ring canal.
22. Note the stone canal connecting the ring canal to the madreporite, where water enters. These canals are difficult to locate without disrupting them, but see if you can identify them.
23. With a magnifying lens, examine the inside wall of the ray to see the supporting ridges, the bulb-like ampullae, tiny sacs that create suction of the tube feet. You may also notice tiny openings in the inner wall.
These pores connect with a fill tube and are part of the external gills that help the starfish to breathe.
24. As with any biological scraps, it is best to contact your local waste management company for proper disposal procedure.
Post-Lab Questions
1. What are some common animal traits that a starfish does not possess?
2. Explain why the starfish is not classified as a vertebrate species.
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Your lab kit will NOT contain the following materials unless specifically requested by your instructor!!!
Experiment 6: Vertebrate Dissection
Frogs are a member of the Amphibia class of vertebrates. In many respects, the anatomy of the frog is similar to human anatomy. Thus, the study of frog anatomy is a useful tool for scientists. As amphibians, frogs may live some of their adult life on land, but return to water to reproduce.
Materials
Dissecting Pins
Flashlight
Dissection Tools
Preserved Grassfrog
Dissection Tray
Ruler
9 cm Fishing Line
Toothpick
Note: To determine the sex of your frog, examine the fingers on its foreleg (arm). A male frog typically exhibits thick pads below the thumbs.
Procedure
1. Place the frog dorsal (back) side up in the dissecting tray. Observe the external anatomy of the skin.
Don’t forget to use your sense of touch in this observation (with gloved hands of course)!
2. Locate the following features:
•
External nares (nostrils)
•
Two tympani (eardrums)
•
Two eyes, each with three lids (the third lid is a transparent covering on the eye)
3. Measure and record the length of the frog in Table 4.
4. Break a toothpick so that you have a two cm piece. Place this in the frog’s mouth to prop it open so you can observe the structures of the frog’s mouth.
5. Using the flashlight, locate the following features:
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•
Internal nares
•
Eustachian tubes
•
Opening of the esophagus
•
Glottis
•
Tongue
Invertebrates and Vertebrates
•
Two kinds of teeth; maxillary teeth help the frog to grip while vornerine teeth point inward.
6. Use forceps to grab the tongue and locate where it attaches to the floor of the mouth.
7. Insert the end of the fishing line into one of the Eustachian tubes and watch the tympanum on the dorsal side of the frog. This system allows air pressure to be equalized in the frog’s head.
8. Place the frog in the dissection tray ventral (belly) side-up, and pin the arms and legs to the tray to stabilize the specimen.
9. Using forceps and dissecting scissors, cut along the midline of the body starting at the cloaca (the urogenital opening) as shown in Figure 14. Make shallow cuts so internal organs are not damaged when cutting through the muscle and breastbone.
10. Make horizontal cuts near the arms and legs, as shown in Figure 14.
Figure 14: The dorsal (above) and ventral (below) sides of the frog. Cut lines are shown on the picture below (green).
11. Pin the flaps to the dissecting tray to expose the internal organs.
Note: If your specimen is female, you may need to remove the eggs and enlarged ovary to view the internal organs.
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12. Using a probe and forceps, lift the internal organs around so you can locate the following organs:
•
Fat Bodies: Located just inside the abdominal wall, these long, thin organs are yellow or orange in color. You may remove these structures if they obstruct your view of underlying organs.
•
Peritoneum: Directly under the body wall, this attached membrane of thin tissue forms a lining around the internal organs. There are various membranes that hold the organs in place, called mesenteries, which are also part of the peritoneum.
•
Liver: A three-lobed organ that sits high in the body cavity. It is the largest organ visible, and is dark brown in color.
•
Heart: A triangular structure that sits above the liver. Note the thin sac (called the pericardial sac) that covers this organ and the vessels extending from it. The pericardial sac can be cut to expose the heart.
•
Lungs: Two spongy organs located beneath the liver.
•
Gall Bladder: This bright green organ can be visualized by spreading apart the lobes of the liver.
•
Esophagus: A tube that transports food from the mouth to the stomach. Insert your probe into the frog’s mouth and observe where it leads.
•
Stomach: Under the left side of the liver, this bag-like digestive organ is the first site of chemical digestion in the frog.
•
Small Intestine: A long coiled tube that serves as a conduit for food and the place where nutrient absorption into the bloodstream takes place. Notice the blood vessels running through this organ.
•
Large Intestine: A widening of the small intestine signals the start of the large intestine. This may be located underneath the small intestines.
•
Spleen: Located in the middle of the body cavity, this dark red, spherical organ stores blood. You may need to lift the stomach and small intestines to see.
13. Using dissecting scissors, remove the stomach from the body cavity. Cut it open to investigate any remains from the frog’s last meal. Record observations in Table 4
14. Remove the small intestines from the body cavity. Measure the length of the small intestines and record in Table 4.
15. Continue to locate the following organs:
•
Kidneys: Dark, bean-shaped organs that filter wastes from the blood. You may have to lift the intestines to see them, as they are towards the back of the body cavity.
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•
Testes (for male specimens): Located above the kidneys, these round organs are typically light in color. •
Oviducts (for female specimens): Curly tubes around the kidneys.
•
Bladder: A bag-like organs that stores urine. Try to trace the tubes from the kidneys to the bladder.
16. As with any biological scraps, it is best to contact your local waste management company for proper disposal procedure.
Table 4: Grassfrog Measurements and Analysis
Frog Length
Small Intestines Length
Stomach Contents
Post-Lab Questions
1. Classify the specimen you just dissected, starting with Kingdom and ending with Species.
2. Describe the appearance of five organs you found in the frog.
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Lab 24
Animal Structure
Animal Structure
Learning Objectives
•
Identify patterns in the organization of organisms including cells, tissues, organs and organ systems
•
Explain how the major structures of the integumentary, skeletal and muscular systems influence their function
•
Compare and contrast the four major types of tissue: epithelial, connective, muscle, and nerve
Introduction
Cells are the fundamental unit of life in every organism. The trillions of cells which comprise the human body are categorized into over 200 different types. Networks of these interconnected cells unite to form tissues capable of performing specialized functions. Although there are over 200 cell types, there are only four tissue types in the human body:
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•
Epithelial tissue - Covers every surface of the body, both internally and externally. Examples include the external layer of skin, as well as the lining of the stomach.
•
Connective tissue - Connects parts of the body. Examples include ligaments and tendons.
•
Muscle tissue - Enables motion and performs specialized functions. Examples include biceps
(move your arm), cardiac muscle (pump blood) and the walls of the digestive tract (force nutrients into bloodstream).
•
Nerve tissue - Provides communication throughout the body. Examples include sensory neurons
(transmit information) and motor neurons (produce impulses to stimulate movement).
Did You Know...
Stem cells are undifferentiated cells that have not yet generated the structures or proteins characteristic of a specialized cell type. Embryonic stem cells are totipotent, meaning they have the potential to become any type of cell in the body (skin, heart, lung, bone, eye, etc.). Adult stem cells are pluripotent, meaning they are already partially specialized. For example, the stem cells in bone marrow can form blood, bone, cartilage and fat, but not a heart.
Adult stem cells are found in relatively small numbers and are thought to maintain and repair tissues. Cell-based therapies induce stem cells to differentiate and repair damaged or destroyed tissues. The goal is for these therapies to be effective in treating conditions such as corneal regeneration, Parkinson’s Disease, Alzheimer’s
Disease, multiple sclerosis, cancer, brain and spinal cord repair, and skin grafts.
Tissue types form organs, which perform specialized functions. Examples of organs in the human body include the heart, kidney, spleen, pancreas, lungs and so forth. Organ systems are comprised of multiple or-
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Animal Structure gans that collectively perform more complex functions. For example, both the bladder and kidneys are a part of the urinary system. There are eleven different organ systems in the human body. The integumentary, skeletal, and muscular systems provide structure and protection to vertebrates.
The Integumentary System
The integumentary system is comprised of skin, nails, hair and some membranes. The skin is the largest component of this system (and largest organ in the body), whose functions include:
•
Protect the body
•
Retain water
•
Host sensory receptors (temperature, pain, and pressure)
•
Maintain body temperature
Hair Shaft
Sweat Pore
Sebaceous gland Arrector pili muscle
Hair follicle
Pacinian corpuscle
Sweat gland
Figure 1: Diagram of skin.
The skin is comprised of three layers:
1. Epidermis - The outermost layer of the skin composed of thick, waterproof cells.
2. Dermis - A more complex layer below the epidermis that houses the sweat glands and hair follicles (hair roots). It has abundant connective tissue which provides elasticity and resilience.
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Animal Structure
3. Subcutaneous Layer - The inner most layer of skin, composed of fat storage cells. It protects and insulates the inner organs.
Figure 2: Skin Layers
The Skeletal System
All animals (both invertebrates and vertebrates) have a support structure that provides protection and allows for movement. Though the systems differ, their functions are similar.
There are three types of skeletal systems:
•
Hydrostatic skeletons - A primitive skeletal system found in soft bodied invertebrates, such as worms or jellyfish, and consists of a fluid filled cavity surrounded by muscle.
•
Exoskeletons - A tough and rigid outer covering, found in many insects.
•
Endoskeletons - A tough and rigid internal support structure that also serves a number of functions (Figure 3), including:
• Provide rigidity
• Provide an attachment site for muscle
• Protect internal organs
Figure 3: The endoskeleton
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Animal Structure
• Allow movement
• Produce blood cells (bone marrow)
• Store minerals (e.g. calcium)
The two primary components of the skeletal system, cartilage and bone, are often connected by tendons (a tough band of connective tissue that join muscle to bone) and ligaments (similar composition to tendons but connect bone to bone). Cartilage is stronger than tendons and ligaments, but is more flexible. It provides a low friction surface and shock absorption for joints.
The primary component of the human skeletal system is bone, which on average accounts for 15% of our total body weight. In addition to providing structure and strength to our body, the inside of bones store bone marrow, which produce a variety of cells critical to normal body function and also repair (e.g. blood cells, fat cells, blood vessels).
There are two types of bone:
•
Compact bone - The hard outer layer of all bones that appears smooth and white. Compact bone comprises about 80% of bone mass •
Spongy bone - A porous structure inside compact bone that houses red bone marrow, supplies nutrients to the bone and produces new blood cells for the body. It is part of most large bones, including the pelvis, skull and vertebrae.
Spongy
Bone
Compact
Bone
Figure 4: Types of bone.
Real World: Arthritis
Arthritis is caused by joint inflammation, and is often seen in the hands, knees, or hips. There are two primary branches of arthritis: rheumatoid arthritis, and osteoarthritis. Rheumatoid arthritis is caused be an autoimmune disorder, whereas osteoarthritis is caused by natural wear and tear experienced by joints in everyday life. Symptoms of both types include joint pain, redness, swelling, and stiffness. The cartilage which coats the ends of the bones connected by the joint, decreases over time and may even result in bone-on-bone grinding in severe cases. As to be expected, osteoarthritic symptoms typically worsen as the body ages. In rheumatoid arthritis, the decrease in cartilage may be accelerated by
Figure 5: Common areas for arthritis. local attacks from the immune system on the lining of the joint capsule.
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Animal Structure
Muscular System:
Muscles enable motion and perform vital functions (e.g., digestion). The approximately 630 muscles in the human body fall into three types and account for 40% of total body mass (on average).
•
Skeletal Muscle: Also known as striated muscle, range considerably in size and shape, have alternating dark and light bands and are connected to bones, enabling motion. Unlike other types of muscle, it is voluntarily controlled. For example, if you want to jump, these muscles contract so you can. Figure 6: Innervation of skeletal muscle (left) and muscle fiber anatomy (right).
•
Smooth Muscle: Found in almost all hollow cavities in the body (e.g., stomach, intestine, bladder, uterus), are smaller than skeletal muscle, do not exhibit striations and are involuntarily controlled
(they function without conscious thought). For example, digestion, which is facilitated by smooth muscle, occurs whether you think about it or not.
•
Cardiac Muscle: Found only in the heart, has alternating striations and is involuntarily controlled.
Cardiac muscle never rests until the heart stops beating.
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Animal Structure
Table 1: Major Systems Matched with Function and Component
System
Function
Components
Nervous
System
Receives and processes information and then directs the body of what to do
Brain, nerves, spinal cord, sensory organs
Endocrine
System
Works with the nervous system, using chemicals (hormones) to communicate with the body and regulate its chemical processes Many glands (pituitary, thyroid, adrenal, etc.), pancreas
Lymphatic
System
Defense system of the body as well as providing transportation of many fluids and compounds
Spleen, lymph nodes, lymphatic vessels, tonsils, thymus
Muscular
System
Moves the body
Skeletal muscle, smooth muscle, cardiac muscle
Skeletal
System
Provides protection and support for the body Bones, ligaments, cartilage, tendons
Digestive
System
Processing and absorption of nutrients from food
Salivary glands, liver, stomach, gallbladder, esophagus, pancreas, teeth, small and large intestines, rectum
Circulatory
System
Transport oxygen, nutrients and other compounds throughout the body and then takes away waste products from the body
Heart, blood, blood vessels
Respiratory
System
Obtains oxygen and transfers waste gases out of the body
Lungs, nose, trachea, diaphragm, pharynx, larynx, bronchi
Urinary System
Excretion of waste (urine) from the body
Kidney, bladder, ureters, urethra
Integumentary
System
Covers and protects the body
Nails, skin, hair, some membranes
Reproductive
System
Prepare cells for and participate in reproduction
Males: penis, testes, seminal vesicles, prostate gland, epididymis, vas deferens
Females: vagina, uterus, fallopian tubes, ovaries 358
Animal Structure
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Did You Know...
Aerobic activity results in a lot more than calorie burn-off! In fact, as soon as you start exercising a biological domino effect takes place. Here’s the breakdown:
1. Your body begins breaking ATP down into ADP and phosphate ions. When these bonds break, energy is released which can be used for muscle contraction.
2. The initial store of ATP is used up within a minute or two. To harness more, glycogen, glucose, and fat molecules are broken down as an alternative energy source to create ATP.
3. Breaking down these molecules requires extra oxygen. To achieve this, heart rate increases and blood is pulled away from unneeded systems like the digestive system. This allows the body to pump more blood quickly, pushing red blood cells throughout the body which drop off oxygen molecules to the muscles.
4. Every time an ATP, glycogen, or glucose molecule is broken down energy is released. Some of this energy is used by the muscles to accommodate the exercise movements, but the energy also results in an increase in body temperature.
5. To maintain a healthy body temperature, the circulatory system comes into play again by focusing blood-flow towards the skin.
This may cause redness, and releases heat as sweat glands are activated to release heat-laden moisture.
6. When a workout is complete, the brain releases a rush of endorphins which contributes to the “runners high” many people feel.
Pre-Lab Questions:
1. Give two examples of how the integumentary system, skeletal system and muscular system interact. 2. Hypothesize how the muscular system works with the circulatory system to maintain homeostasis.
3. Relate tissues to organs and organ systems. Give an example.
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Experiment 1: Examining Skin, Bones and Muscle Histology
The skin (the primary component of the integumentary system) is a large organ which provides protection to the body. Muscles and bones are located beneath the skin and enable movement, structure, and further body protection. In this experiment you will observe the structure and function of cells of the integumentary, skeletal, and muscular systems.
Materials
Skin Digital Slide Images
Cardiac Muscle Digital Slide Image
Cortical (Compact) Bone Digital Slide Image
Skeletal Muscle Digital Slide Image
Trabecular (Spongy) Bone Digital Slide Image
Smooth Muscle Digital Slide Image
Note: All images are located at the end of the procedure.
Procedure
Part 1: The Skin
1. Observe the digital slide picture of the hairy animal at 100X, 400X, and 1000X.
2. Note the locations of the epidermis, dermis, dermal papillae, and the sweat glands. Note that fat cells that comprise the subcutaneous layer.
Part 2: Investigating Cortical (Compact) vs. Trabecular (Spongy) Bone
1. Examine the digital slide images of the cortical (also referred to as compact) and trabecular (also referred to as spongy).
2. Be sure to read through and locate all of the labeled features.
Part 3: Muscle Structure
1. Examine the digital slide images of cardiac, skeletal, and smooth muscle.
2. Again, review all of the labels and annotations presented with each image.
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Animal Structure
Human Skin 100X.
Epidermis
Layer
Sweat gland duct
Dermal Papillae
Dermis Layer
Human Skin 400X. The stratum corneum contains primarily keratinized cells. Dermal papillae can form ridges that aid in gripping (fingerprints).
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Animal Structure
Human Skin - A 1000X. The layers of the epidermis can be distinguished. The keratinocytes of the stratum granulosum contain dark staining granules.
Hair follicle
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Human Skin- B 1000X. Enlargement of a hair follicle located in the dermis. The keratinized hair root and the hair matrix (region of cell division) are enclosed by the hair follicle.
Animal Structure
Haversian system Haversian canals
Interstitial lamellae
Concentric lamellae Cortical Bone (Compact bone) 100X. Cylindrical units called osteons are notable in cortical bone.
Lacunae containing osteocytes
Canaliculi
Lamella
Cortical Bone 1000X. Osteocytes, mature bone cells, are located in cavities called lacunae. Canaliculi provide passageways for nutrients traveling to the osteocytes.
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Animal Structure
Trabeculae
Ma
r
ro w Ca v it y
Trabecular Bone (Spongy Bone) 100X. Marrow fills the internal cavities found in trabecular bone.
Trabeculae
Marrow Cavity
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Trabecular Bone 1000X.
Animal Structure
Branched structure
Cardiac Muscle – A Tissue 100X. Note the branched structure.
Cardia
c mus cle cell nu cleus Inte r cala ted di
scs
Cardiac Muscle Tissue 1000X.
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Animal Structure
Cardiac Muscle Tissue – B 100X. Cardiac muscle is well known for its location and striations. It is only found as a component of the heart walls. They are branched in structure and possess one nucleus (as opposed to skeletal muscle which is cylindrical in shape and multi-nucleated). Cardiac muscle also possess intercalated discs which hold the cardiac muscle fibers together when the heart undergoes contraction.
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Animal Structure
Muscle Fibers
Skeletal Muscle 100X. The multinucleated skeletal muscle fibers run parallel to each other.
Nuclei
Single Muscle
Fiber
Skeletal Muscle 1000X. Striations resulting from the arrangement of contractile proteins that run perpendicular to the muscle fiber.
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Animal Structure
Smooth Muscle Tissue 100X. Smooth muscle tissue surrounds the gastrointestinal tract and helps. Similarly, it also helps blood vessels contract and push blood through the circulatory system.
Smooth Muscle Tissue 1000X. These cells can be described as spindle-shaped.
Note the elongated ends of the cells.
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Animal Structure
Post-Lab Questions
1. Label the arrows in the following slide image:
A
B
C
Human Skin 400X.
2. Which layer comprises the majority of skin?
D
Explain how you came to your conclusion using your
knowledge of skin components and functions.
3. What component of the subcutaneous layer provides protection to the underlying organs?
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Animal Structure
4. What layer of skin causes dandruff? Is this the same layer that peels off after a sunburn? How do you know? 5. Wrinkles occur with age due to a loss of elasticity and collagen. Which layer of skin do you believe is most responsible? Do you think topical ointments that claim to firm skin really work? Explain why or why not? 6. Sketch a drawing of a bone (you pick!), and label the compact and spongy regions.
7. Identify at least two differences and two similarities between cortical and trabecular bone. These may be anatomical or physiological.
8. Why do you think spongy bone allows more flexibility than compact bone?
9. Suppose you are looking at an unlabeled slide image of muscle. List two items which would help you determine if the muscle was cardiac, smooth or skeletal?
10. Why does muscle account for such a large portion of your body mass? Explain using scientific evidence.
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Animal Structure
Experiment 2: Calcium in Bones
As a child, you may have been encouraged to drink milk so that “your bones would grow strong.” Milk is high in calcium, one of the primary components of bones. On average, 99% of all of your body’s calcium is stored in bones. If your body becomes calcium deficient, it will use the reserve stored within your bones.
Ultimately, your bones can lose their strength if this reserve is depleted.
Remarkably, bones are fairly light weight but can resist large amounts of stress. In addition to calcium, collagen fibers provide strength and flexibility, enabling bones to withstand physical pressure, resistance, impact, tec. If these components are damaged, the structure of the bone may also change.
In the following experiment you will examine the importance of calcium in bones. Egg shells possess calcium, and acetic acid (vinegar) removes calcium from bone.
Materials
Figure 3: Endoskeleton
200 mL Acetic Acid (Vinegar), C2H4O2
Permanent Marker
Aluminum Foil
*Water
(2) 250 mL Beakers
Construction Paper (Black)
*You Must Provide
*1 Egg
Note: This lab requires five days of observation. Wear gloves when handling egg shells.
Procedure
1. Use the permanent marker to label the two 250 mL beakers as 1 and 2.
2. Measure and pour 200 mL of water into Beaker 1.
3. Measure and pour 200 mL of acetic acid into Beaker 2.
4. Carefully crack one egg into two, approximately equal sizes. Discard the yolk and gently rinse the egg shells under running water.
5. Use a gloved hand to gently place the first half of the egg shell in Beaker 1. If possible, try to submerge the egg shell by filling any vacant cavities with water. Cover the beaker with aluminum foil and let sit at room temperature for five days.
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Animal Structure
6. Use a gloved hand to gently place the second half of the egg shell in Beaker 2. Again, try to submerge the egg shell. Cover the beaker with aluminum foil let sit at room temperature for five days.
7. Divide the black construction paper in half. Label one side as water and one side as acetic acid.
8. On the 5th day, remove the acetic acid and water from Beaker 1 and Beaker 2, respectively.
9. Use a gloved hand to remove each of the egg shells from the beakers, and place them one the corresponding side of the construction paper.
10. Visually examine the physical characteristics of the egg shells. Record your observations in Table 2.
11. Use your fingers to gently apply pressure to the egg shells. Notice the differences in the strength or flexibility. Record your observations in Table 2.
Table 2: Effect of Acetic Acid on Egg Shells
Beaker
Observations
Beaker 1: Water
Beaker 2: Acetic
Acid
Post-Lab Questions
1. What do these results suggest about the role of calcium in bones? Is it important? Why or why not?
2. Why did the flexibility of the egg shell change after it was removed from the acetic acid?
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Animal Structure
Experiment 3: Agonists and Antagonists
Many muscles work in antagonistic pairs; meaning, when one muscle contracts, another relaxes. For example, when the bicep contracts it lifts the forearm up and lengthens and at the same time. The forearm is forced down and the bicep lengthens (causing it to relax) when the tricep contracts.
There are two primary types of muscle contractions: isotonic and isometric. Isotonic contractions occur when the length of a muscle changes. Isometric contractions do not change the muscle length.
Materials
Figure 3: Endoskeleton
Figure 7 (located at end of procedure)
Procedure
Part 1
1. Identify and mark as many antagonistic pairs as possible for the following body regions (hint, it will help to feel the muscles on your own body to determine the antagonistic pairs):
A. Lower leg
B. Upper leg
C. Lower arm
D. Upper arm
Record your pairings in Table 3.
2. Lay your left arm on a flat surface with the palm up. Place your right palm on top of your left bicep and slowly raise your left forearm. Notice the shortening of the bicep muscle. That is an isotonic contraction.
3. Stand, hold your left arm in front of you and slowly squeeze your fist. Notice that the muscles in the forearm do not shorten when contracted. That is an isometric contraction.
Part 2
4. For the following exercises, perform the muscle contraction and determine if it is an isotonic or isometric contraction. a. Place your hand underneath the table. Push up on the table up while keeping your arm straight.
b. Lay on your back on the floor. Pull your chest up to your knees (a sit up).
c. Sit in a chair and place a ball (or other object) between your feet. Slowly lift your feet into the air.
d. Lie on your side on the floor and raise your upper leg towards the ceiling.
e. Sit in a non-moveable chair and place your feet straight out in front of you against a solid object
(like your desk). Try to push away the desk.
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Animal Structure
Figure 7: Human Muscle
Table 3: Antagonistic Muscle Pairings
Body Region
Lower Leg
Upper Leg
Lower Arm
Upper Arm
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Paired Muscles
Lab 25
The Circulatory and Respiratory Systems
The Circulatory and Respiratory Systems
Learning Objectives
•
Explain the function of the heart and blood in the circulatory system
•
Describe the structure and function of the respiratory system highlighting respiration
•
Explain how the circulatory and respiratory systems work together to maintain homeostasis
Introduction
All cells and ultimately systems, in the human body require nutrients and oxygen. They must also be capable of eliminating accumulated waste products. The circulatory and respiratory systems play a major role in these processes.
The Heart
The circulatory system is driven by the heart. As the heart contracts, blood is propelled from the left ventricle into the aorta. From there, it travels through successively smaller arteries before reaching arterioles, capillary beds, and finally tissues. Together with the heart, the circulatory system is responsible for distributing blood and nutrients to all the cells of the body.
Blood
Figure 1: The heart is divided into two chambers containing four regions: right atrium, right ventricle, left atrium, and left ventricle.
Blood, a fluid tissue, is a mixture of plasma, red blood cells, white blood cells, and platelets (Figure 2). The blood that circulates throughout the veins and arteries of the body is made of a variety of cells and cell fragments suspended in a liquid called plasma. Plasma makes up the largest portion of blood, accounting for 55% of the fluid. Plasma itself is nearly 90% water. Table 1 lists different types of cells found in blood and their corresponding functions.
Blood serves three main functions:
1. Transport oxygen, carbon dioxide, nutrients, hormones, and waste.
2. Regulate pH, body temperature, and water content of cells.
Figure 2: Red and white blood cells attacking a pathogen.
3. Protect against blood loss (through clotting) and disease through phagocytic white blood cells and antibodies.
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The Circulatory and Respiratory Systems
Table 1: Types of Blood Cells
Erythrocytes
Also known as red blood cells; comprise approximately 45% of blood. Contain hemoglobin for oxygen and carbon dioxide transport.
Lymphocytes
Involved in the immune response; produce antibodies.
Monocytes
Thrombocytes
Phagocytotic macrophages.
Also known as platelets. Involved in blood clotting.
Neutrophils
Key to wound healing.
Eosinophils
Phagocytosis.
Basophils
Leukocytes
Release histamines.
Also known as white blood cells; comprise approximately 1% of blood. Involved in fighting disease and foreign objects in the body as part of the immune system.
Red blood cells have special regions that bind and transport respiratory gases (O2 and CO2). This exchange of gases and nutrients is known as respiration. White blood cells are one component of the body’s defense system that attack foreign organisms or particulates including bacteria and viruses. They also produce antibodies. Platelets are involved responsible for clotting blood.
The Circulatory System
The circulatory system is a closed system where the blood remains in the vessels (arteries and veins) and does not flow freely in the organs. Blood flows throughout two circuits, both of which provide a mechanism for gas exchange: the pulmonary and the systemic (Figure 3).
The pulmonary system adds oxygen (O2) to the blood and removes carbon dioxide (CO2). Alveoli are small nodules located on deep braches of the lung that have a large surface to volume ratio. The gas exchange takes place between these nodules and the hemoglobin, a protein in red blood cells. The systemic system delivers oxygenated blood to the cells through capillaries and removes waste products, including CO2. All cells reside within close proximity to capillaries to facilitate this process. Both systems are driven by the heart.
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Figure 3: The circulatory system showing the pulmonary system in red and the systemic system in blue.
The Circulatory and Respiratory Systems
Oxygen depleted blood travels through the vena cava into the right atrium. The right atrium contracts, forcing the blood into the right ventricle. The right ventricle contracts, pushing the blood into the pulmonary vein where it travels to the lungs and exchanges CO2 for O2. The blood then returns to the left ventricle.
The ventricle contracts, ejecting the blood into the left atrium. The left atrium contracts and pumps blood into the aorta, which delivers it to the rest of the body. A pulse is created when blood is pumped by the heart into the aorta (systolic pressure). Blood pressure on the vessels (when heart muscle relaxes) creates the diastolic pressure. These two measurements are taken as your blood pressure in a doctor’s office to monitor the health of your circulatory system.
Figure 4: Anatomy of the heart.
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The Circulatory and Respiratory Systems
The blood also makes its way to the digestive system where it picks up nutrients and to the urinary system, where it disposes of waste products. Together, the respiratory system and circulatory system provide the body with oxygen, and a delivery route for nutrients.
Cells require oxygen to produce ATP by cellular respiration. The process produces CO2 as a byproduct, a toxin that must be removed. The respiratory process relies on the diffusion of gases (O2 in and CO2 out) across the cell membranes (Figure 4).
The Respiratory System
The main organ of the respiratory system is the lung. This spongy, elastic tissue stretches and contracts as you breathe. Air travels through your nose or mouth to the trachea, which branches into many smaller tubes called bronchi and then bronchioles. This airway leads to microscopic air sacs called alveoli, which are covered with capillaries. This is the first site of gas exchange.
Respiration involves three steps to transport the oxygen (with the help of the cardiovascular system) and carbon dioxide throughout the body.
Pulmonary ventilation is the movement of air between the atmosphere and the lungs. It occurs during inhalation and exhalation. The diaphragm and the muscles of the thoracic wall change the space and pressure inside the lungs. When the diaphragm contracts, it leaves more space for the lungs to expand and lowers the internal air pressure. Air pressure is greater outside of the body, and will move to a region of lower pressure (inside the lungs). External respiration is the movement of oxygen from the alveoli into pulmonary capillaries and carbon dioxide from the pulmonary capillaries to the alveoli. Oxygen and carbon dioxide are transported between the lungs and body tissues. Internal respiration is the movement of oxygen from the capillaries into body cells, and of carbon dioxide from the body cells into ca- Figure 5: The trachea subdivides into pillaries. bronchi, bronchioles, and smaller
The steps involved in respiration are:
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•
Ventilation
•
Gas exchange between alveolar air and lung capillaries
•
Bulk flow of the circulation
•
Gas exchange between the capillaries and tissue cells
•
Cellular consumption and production of gases
conduits that conduct air from the external environment to the alveoli, the site where gas exchange occurs.
The Circulatory and Respiratory Systems
Molecules of oxygen and carbon dioxide are passively exchanged through the process of diffusion into and out of the blood. Diffusion is the net movement of molecules from a region of higher concentration to a region of lower concentration as a result of their random movement.
Figure 6: Gas exchange: oxygen (red) diffuses from the lungs to the blood stream and is used in aerobic respiration. Carbon dioxide (green and yellow), a waste product of respiration, diffuses from the bloodstream to the lungs where it is exhaled out of the body.
Pre-Lab Questions
1. Identify one reason why carbon dioxide is considered a waste product to the body.
2. How does pressure in the lungs change when a person inhales? How does it change when a person exhales?
3. Explain the role of hemoglobin in the circulatory system in 1-2 sentences.
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The Circulatory and Respiratory Systems
Experiment 1: Understanding Lung Mechanics
In this experiment, you will model lung function.
Materials
Balloon
Wash Bottle
Rubber Band
Procedure
1. Attach the balloon to the end of the straw that fits inside the wash bottle, and secure with a rubber band.
The balloon will act as the lung.
2. While gently squeezing the bottle, replace the top and secure tightly.
3. Squeeze the bottle and record what happens to the balloon in Table 2.
4. Release the bottle and observe what happens in Table 1.
Table 1: Understanding Lung Mechanics Observations
Squeezed Bottle Observations (Step 3)
Released Bottle Observations (Step 4)
Post-Lab Questions
1. What happens to the lung (balloon)? Why?
2. What would happen if the seal at the base of the bottle leaked?
3. What causes a collapsed lung?
4. Is a collapsed lung functional? Why or why not?
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Experiment 2: Observation of Blood
Materials
Digital Slide Images of Blood
Procedure
1. Examine the digital slide picture of blood. Then answer the Post-Lab Questions.
Blood Smear 100X
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The Circulatory and Respiratory Systems
Blood smear 1000X. Blood cell types shown include erythrocytes (red blood cells), lymphocytes (the sparse, large, pale pink circles), and neutrophils.
Typical red blood cells
Sickled red blood cells
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Red Blood Cells 1000X. Red blood cells (RBCs) are typically circular in shape. Patients with sickle cell anemia produce blood cells with atypical cell shapes; these tend to appear elongated or suckered near the middle of the cell.
The Circulatory and Respiratory Systems
Post-Lab Questions
1. What is the most prevalent cell type in blood? Hypothesize why that is.
2. Based on what you have learned, how would you explain carbon monoxide poisoning?
Experiment 3: Modeling the Circulatory System
In this experiment you will explore and model the structures and function of the circulatory system.
Materials
2 Check Valves
60 cm of Tubing
2 Drops of Dye
*Scissors
Nail
*Water
Permanent Marker
2 Plastic Bottles
*You Must Provide
Procedure
1. Use the permanent marker to label the plastic bottles 1 and 2.
2. Use a nail to carefully puncture two holes in the side of each bottle (approximately 3.5 cm from the bottom of the bottle) so that the holes are across from each other and at the same height from the bottom of the bottle. It is best to puncture the bottle with the nail straight through both sides. This will help keep the holes level.
LAB SAFETY: BE EXTREMELY CAREFUL WHEN PUNCTURING THE BOTTLE. KEEP IN MIND
WHERE YOUR HANDS ARE AT ALL TIMES.
3. You may need to maneuver the nail around the hole to make it large enough to fit the plastic tubing
(Figure 7). The goal is for the tubing to fit tightly in the hole.
4. Carefully cut the 60 cm tubing into four equal lengths of 15 cm.
5. Attach one end of one piece of 15 cm tubing to the OUT end of a check valve.
6. Attach one end of a second piece of tubing to the IN end of the check valve.
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The Circulatory and Respiratory Systems
Figure 7: Maneuver the nail in a circular motion to enlarge the hole so that the tubing fits tightly.
7. Repeat Steps 3 and 4 for the remaining check valve.
8. Construct a closed system by inserting the free ends of the tubes into the holes you created in the plastic bottles to create a loop. The tubes should be far enough into the bottle so that water can flow from tube to bottle to tube. The tubes in each bottle should not touch (Figure 8).
Note: Properly assembled models should have the direction of the check valves facing in opposite directions.
9. Fill each bottle ¾ with water. The water level should be higher than Figure 8: Model Set Up the tubing.
10. Place the model on an underpad or paper towel to protect surfaces for the next step. This will also help catch water that leaks from the model. There should be minimal leaking if tubes are tightly in the bottles.
11. Place one drop of dye in each bottle and close the bottle by screwing on the caps.
12. Gently swirl the bottles to allow dye and water to mix. Be careful not to squeeze the bottles!
13. Note where the water is initially before squeezing the bottles and record your observations in Table 2.
14. Squeeze bottle 1 with both hands. Observe the direction of water flow and the water level in each bottle while you are squeezing and right after you stop squeezing. Record your observations in Table 2.
15. Repeat Step 13 for the bottle 2 and record your observations.
16. Squeeze one bottle and then the other back and forth. Observe the direction of water flow and the water level in each bottle. Record your observation in Table 2.
17. Draw a diagram of your model under Table 2.
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The Circulatory and Respiratory Systems
18. Use the terms Left Side of Heart, Right Side of Heart, Lungs and Body and knowledge gained from the
Introduction to label your drawing.
19. Use arrows to label your drawing to show the direction of water flow.
Table 2: Circulatory System Model Observations
Action
Water Level and Direction of Water Flow
Before Squeezing the Bottles
Squeezing Bottle 1
Directly After Squeezing Bottle 1
Squeezing Bottle 2
Directly After Squeezing Bottle 2
Squeezing Bottle 1 and 2 Back and Forth
Diagram of Model
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The Circulatory and Respiratory Systems
Post-Lab Questions
1. What did each of the bottles represent? What does the water represent?
2. What circulatory system structure do the check valves represent? What is their function in the circulatory system?
3. What do you predict would have happened if there were no valves present in the model? How would this affect the human body?
4. Where on the model did you assign the Lung and Body labels? Use circulatory system structures when answering.
5. Why use two bottles to represent the heart instead of just one? Use scientific evidence to support your answer. 6. Humans have a four-chambered heart. How could you modify the model to show this? What supplies would you need? Draw a diagram of a possible model.
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Lab 26
The Sensory and Nervous Systems
The Sensory and Nervous Systems
Learning Objectives
•
Identify the structures and function of neurons including dendrites, axons and synapses
•
Identify the major structures and functions of the central and peripheral systems
•
Distinguish between the somatic and autonomic systems of the peripheral nervous system
Introduction
The nervous system carries a large responsibility for maintaining the homeostasis within the body. It is the master control and communication center of the body. The nervous system allows the body to interpret, integrate, and react to the surrounding environment as well as the internal stimuli occurring simultaneously throughout the body. Voluntary actions such as writing, reading, or running, as well as involuntary actions such as the heart pumping blood through the body, the pancreases releasing enzymes, or the gut digesting food are monitored by this complex system.
The three main responsibilities of the nervous system are sensory input (for stimuli inside and external to the body), integration of sensory input, and a motor response by activating effector organs. To accomplish these substantial tasks, the nervous system is organized into two distinct sub-systems: the central nervous system (CNS) and the peripheral nervous system (PNS).
Special cells called neurons transmit signals to and from the brain, while surrounding glial cells maintain the neurons so they operate at their peak performance. Each neuron has a long arm called the axon, which sends signals to other neurons’ dendrites
(Figure 2). What are commonly called ‘nerves’ are really the axonal processes of nerve cells. These processes are insulated with myelin to ensure signals are sent quickly.
It is important to remember that many neurons throughout the body work together to transmit signals from the brain to tissues, and from tissues back to the brain. The junction of two neurons is called a synapse (synapse), and is the site of biochemical signal
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Figure 1: Nervous system anatomy.
The Sensory and Nervous Systems
(neurotransmitter) exchange. Many drugs target synapses and the molecules that exchange between synapses to regulate behavior.
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Did You Know...
There are as many as
1,000,000,000,000,000 synapThere are two parts to the nervous system: the central nervous system and ses in the human brain! Thethe peripheral nervous system. The central nervous system includes the se provide the power to probrain and spinal cord, and is responsible for most of the information pro- cess all of the information our brain receives from the cessing, storage, and retrieval. nerves all over the body.
The peripheral nervous system consists of all the neurons outside of the brain and spinal cord, and can be further classified by function.
•
The somatic nervous system coordinates the body’s movements and for processing external stimuli. This division of the nervous system is regulated by conscious control, and is used for activities such as running, swimming, brushing your teeth, etc.
•
The autonomic nervous system is responsible for the unconscious control of the body, and is further separated into three divisions:
The sympathetic nervous system responds to impending danger or stress, and is implicated in the “fight or flight” response. It is also responsible for control of certain activities that people never think about having to do, such as breathing, blood pressure, and heart rate.
The parasympathetic nervous system is responsible for changes observed during relaxation, such as the slowing of the heart beat, relaxation of pupils, and stimulation of digestion.
The enteric nervous system is responsible solely for digestion. This includes every action from the time food enters the mouth until it exits the body.
Motor neurons transmit messages from the spinal cord to muscles. They project beyond the spinal cord and stimulate muscles by innervating them. Sensory nerve cells transduce information about physical or chemical stimuli to the nervous system for interpretation by the brain.
The Five Senses
The five senses recognized by humans are:
1
Sight
2
Sound
3
Smell
4
Touch
5
Taste.
Figure 2: Neuron.
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The Sensory and Nervous Systems
When these senses are experienced, sensory cells signal the neurons, which transmit the information to the brain via motor neurons. The brain processes the signal and directs the effectors, such as muscles and glands. Some of these responses are voluntary, while others, such as reflexes, are involuntary.
Sensory receptors not only perceive the type of stimulus (temperature, presDid You Know... sure, sight, taste, sound), but also the intensity, location, and duration. Different receptors receive different types of stimuli and patterns that provide addi- Multiple sclerosis is a disease tional information to the brain. in which the myelin coating
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Another interesting aspect of the sensory system is vision. What is sight at the cellular level? A ray of light hits the object being viewed, then reflects into the eye through the lens. Then the light (the image) hits the retina, which has receptors that signal to the brain what color the ray is. Then, the brain receives this information and assembles the colored rays of light into an image.
on neurons deteriorates.
Without the myelin covering, the bioelectric messages sent through the neurons don’t travel far enough to transmit messages to other cells.
The retina has two kinds of receptors – rods and cones. Rods receive black and white rays, while cones detect colors. There are three kinds of cones: red, blue, and yellow. All the colors that are seen are made up of combinations from these three colors. A phenomenon called after image occurs when selected rods or cones tire out and stop working momentarily. When this occurs, the color seen is the a combination of what colors the remaining rods and cones see (usually if you stare at a blue object you will see yellow, if you stare at red you will see green, and vice versa).
4
1 = Aqueous Humor
2
3
2 =Vitreous Humor
1
3 = Sclera
4 = Choroid
10 7 6
9
4
5 = Ciliary Body
6 = Iris
7= Pupil
8
5
8 = Retina
9 = Optic Nerve
10 = Lens
Figure 3: Eye anatomy.
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The Sensory and Nervous Systems
Signals from the brain instruct the rest of the body on how to adapt to stimuli. For instance, in order to see objects at different distances, muscles in the eye contract and relax in order to change the shape of the eye to focus. When looking at a distant object, the lens in the eye is relatively flat. However, the lens becomes more round when looking at a closer object.
The brain receives large amounts of information from our surroundings via sensory organs. Sensory stimuli are detected, processed, and translated into another action through the nervous and sensory systems. The body is an amazing machine, with intricate internal systems that allow us to perceive our environment in multiple ways, and helps us to react to it accordingly in learned or automatic (genetically programmed) responses.
Pre-Lab Questions
1. Explain how the nervous system works with each of the other human body systems.
2. Hypothesize why some human body functions like breathing and heartbeat are an unconscious, involuntary effort.
3. Relate dendrites to axons and synapses.
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The Sensory and Nervous Systems
Experiment 1: Observation of the Nervous System
In this experiment you observe the structures of the neuron and hypothesize how this determines its function.
Materials
Nervous System Digital Slide Images
Procedure
1. Examine the digital slide pictures of neurons. Be sure to review the labels and annotations accompanying each image.
Axon
Cell Bodies
Spinal Cord Smear 100X. Neuroglial cells are closely associated with neurons as they act as supporting cells. 396
The Sensory and Nervous Systems
Neuroglial
Cells
Dendrites
Cell Body
Spinal Cord Smear 1000X.
Nerve fiber
Perineurium
Myelinated Nerve Fiber 100X.
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The Sensory and Nervous Systems
Myelinated Nerve Fiber 1000X. Schwann cells create a myelin sheath by wrapping around the fiber in a ‘jelly- roll’ fashion.
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The Sensory and Nervous Systems
Post-Lab Questions
1. Label the arrows for A and B in the slime image, based on your observations from the experiment:
A
B
2. Hypothesize what the neuroglial cells function is. Support your hypothesis using scientific evidence.
3. Provide a hypothesis explaining how the axon and the dendrites affect the function of a neuron.
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The Sensory and Nervous Systems
Experiment 2: Limits and Capabilities of the Eye
In this experiment you will explore the visual component of the sensory system.
Materials
Image of Blue Triangle (located at the end of the procedure) Image of Yellow Triangle (located at the end of the procedure) Ruler
*Pencil/Pen
*White Construction or Printer Paper
* You Must Provide
Procedure
Part 1
1. Hold the pencil by the eraser with the point facing the ceiling. It should be at arm’s length in front of your right eye.
2. Close your left eye, and slowly move the pencil toward your eye.
3. Focus on the end of the pencil.
4. Keep moving the pencil until the tip is out of focus. Measure the distance (called the near point) between your eye and the pencil in cm. Record your measurement in Table 1.
5. Repeat Steps 2 - 4, this time closing your right eye. Record your measurement in Table 1.
6. If you wear glasses, repeat these steps for each eye with glasses off. Record your measurements for the right and left eyes in Table 1.
Part 2
7. Quickly review the yellow and blue triangles (located at the end of this procedure).
8. Formulate a hypothesis stating what you think will happen if you stare at the yellow triangle for 30 seconds. Then, create a second hypothesis stating what you think will happen if you stare at the blue triangle for 30 seconds. Record your hypothesis in Post-Lab Question 1.
9. Stare intently at the yellow triangle for 30 seconds. Quickly shift your gaze to a sheet of white paper.
10. Repeat Step 9 using the blue triangle.
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The Sensory and Nervous Systems
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The Sensory and Nervous Systems
Table 1: Left and Right Eye Measurements
Eye
Near Point (cm)
Near Point (without glasses, if applicable; cm)
Right Eye
Left Eye
Post-Lab Questions
1. Include your hypotheses from Part 2: Step 7.
2. Examine Table 2. According to the table, how “old” are your eyes?
Table 2: Age Point Correlation Table
Age (years)
10
20
30
40
50
Near Point (cm)
9
10
13
18
50
3. Identify the two primary fluids found within the eye. Which one of these fluids provides nutrients to the rest of the eye? Hypothesize why this is important.
4. What did you see after you stared at the yellow triangle, and then looked at the white paper? What happened when you looked at the blue triangle?
5. Use scientific reasoning to explain what is happening when you stare at the image, then transfer your gaze to a blank sheet of paper.
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The Sensory and Nervous Systems
Experiment 3: Skin Response
In this experiment you will explore how the nervous system interacts with the integumentary system.
Materials
(3) 250 mL Beakers
*Pencil/Pen
16 Paperclips
*Participant (not the student)
Ruler
*Tape
Toothpick
*Warm Water
*Ice Water
*Room Temperature Water
* You Must Provide
Note: You will need a laboratory partner to perform Steps 1 - 4 of this experiment.
Procedure
Part 1
1. Make a caliper by unwinding two of the paperclips until they are straightened. Then, bend each paperclip in the middle to form a U-shape. There should be approximately 0.5 cm between the two tips of each paperclip after being bent into a Ushape.
2. Tightly wrap each tip with a small piece of adhesive tape. This
Figure 4: Paperclip shape reference.
minimizes any sharp points on the paperclip. See Figure 4.
3. Repeat Steps 1 - 2 for the remaining paperclips with the ends set 1.0, 1.5, 2.0, 2.5, 3.0, 3.5 and 4.0 cm apart. When finished, you should have two calipers for each other the eight possible sizes.
4. To test the sensitivity of different body regions, ask your partner to close his/her eyes. Start with the largest caliper (the one with 4.0 cm between the tips) and place both tips of the paperclip onto the left side of your partner’s scalp. Do NOT tell your partner if you place one or both of the tips on his/her head.
5. Ask your partner how many points s/he can distinguish. If s/he can feel two points, move on to the 3.5 cm caliper and test again (your partner should keep his/her eyes closed!). Repeat until your partner can only feel one point. Record the caliper size at which only one point can be felt in Table 3.
6. Continue to test different regions on the left side of the body. Use Table 3 to select body regions.
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7. Repeat Steps 4 - 5 on the right side. Observe if there is a difference between the left and right sides of the body, and record your results in Table 3.
Part 2
8. Obtain three beakers. Fill beaker one with ice water, beaker two with room temperature water, and beaker three with warm water. Immerse your left pointer finger in the ice water, and your right pointer finger in the warm water for 30 seconds.
9. Then, place the finger that was in the ice water into the beaker filled with room temperature water. Note the sensation (did it feel cooler or warmer?) in Table 4.
10. Next, place the finger that was in the warm water into the room temperature water. Again, note the sensation in Table 4.
Table 3: Two-Point Discrimination Test Results
Body Region
Scalp
Forehead
Lips
Front of Neck
Back of Neck
Shoulder
Upper Arm
Elbow
Forearm
Wrist
Back of Hand
Palm of Hand
Tip of Thumb
Tip of Second Finger
Tip of Third Finger
Tip of Fourth Finger
Tip of Fifth Finger
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Left Side Caliper Measurement
Right Side Caliper Measurement
The Sensory and Nervous Systems
Table 4: Temperature Sensation
Initial Beaker/Final Beaker
Sensation
Ice Water/Room Temperature Water
Room Temperature Water/Ice Water
Post-Lab Questions
1. Which of these areas (forearm, back of the neck, index finger, and the back of the hand) contains the most touch receptors? Why is this useful?
2. What was the sensation for each hand when you placed them in the room temperature water?
Experiment 4: Brain Mapping
The brain is the control center of the body. Different areas of this organ are responsible for different functions. In the following exercise, you will learn the anatomy of the brain and the functional areas important to our everyday actions.
Materials
Figure 6
Permanent Marker
**White Swim Cap
**If you have a latex allergy, wear vinyl gloves when drawing on the cap, and do not put it on your head! Note: Feel free to use as many additional marker colors as you like.
Procedure
1. Study Figures 5, 6, and 7 noting the major functional areas of the brain.
2. Lay the swim cap on a flat surface, or you may choose to ask a volunteer to put it on while you map!
3. On the left side of the swim cap, outline the following structures:
Cerebral hemispheres (left and right)
Cerebellum
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The Sensory and Nervous Systems
Central sulcus (divides parietal and frontal lobes)
Lateral sulcus (divides the front and parietal lobes)
4. Outline and label the following lobes:
Frontal
Temporal
Parietal
Occipital
5. Number the functional areas of the cortex located in the lobes, using the number key on Figures 7 and 9
(lateral and superior views only).
6. Wear your cap and explain to someone what each of the functional areas you labeled does!
10
1 = Frontal Lobe
11
2 = Parietal Lobe
2
5
1
3 = Occipital Lobe
12
4 = Temporal Lobe
6
8 9
5 = Brocca’s Area
6 = Tertia Optica Area
7
7 = Wernicke’s Area
3
4
8 = Pars Triangularis
9 = Pars Opercularis
14
13
10 = Gyrus Praemotoricus
11 = Gyrus Praecentralis
Figure 5: Brain anatomy (lateral view).
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The Sensory and Nervous Systems
Figure 6: Sagittal plane.
Broca’s Area
Touch
Corpus Callosum
Writing
(right-handed)
Auditory Cortex
(right-ear)
Auditory Cortex (leftear)
Spatial Visualization and
Analysis
Wernicke’s Area
(language,
math, etc.)
Visual Cortex
(right-field)
Visual Cortex (left
-field)
Figure 7: Superior view. The right and left cerebral hemispheres are connected by the corpus
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callosum, allowing them to coordinate activity.
The Sensory and Nervous Systems
Post-Lab Questions
1. Describe the function of three areas of the brain.
2. Hypothesize why major functions of the human body are regulated by specialized parts of the brain.
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Lab 27
Ecology of Organisms
Ecology of Organisms
Learning Objectives
•
Define ecology as interactions amongst organisms and their environment
•
Analyze the effects of abiotic and biotic factors in an ecosystem
•
Assess how generalists and specialists define habit tolerance
Introduction
Ecology is the scientific study of both the interactions among organisms and the interactions between organisms and their environment. It is an interdisciplinary field that blends Earth science, evolutionary biology and general biology. Ecology explores the adaptations of organisms, the distribution of organisms and biodiversity.
Group of communi es
Group of popula ons
Group of conspecific organisms
Individual
Ecology can be divided into different levels or categories. Ecoform of life system ecology looks at the “big picture”. It focuses on the cycling of chemicals and the energy flow within an ecosystem. EcoTissues or systems system ecology includes all the abiotic factors and species within an ecosystem. Community ecology is concerned with the interSmallest actions between species and how those interactions affect the unit of life structure and organization of the community. As the name implies, population ecology is concerned with populations. A popu- Figure 1: Note the relationship between lation is a group of individuals of the same species living in the species, populations, communities and ecosystems. same area. Population ecology focuses on factors that affect population density and growth. Ecology of organisms, or organismal ecology, is the study of individual organisms and the adaptations that have enabled them to survive in their abiotic environments.
Abiotic and Biotic Factors
Environments encompass abiotic and biotic components. Abiotic components include the physical and chemical aspects. Physical factors that affect an ecosystem include sunlight, temperature, water currents, wind, nature of soil, latitude and altitude. Chemical factors include available nutrients (in soil or water), water salinity, toxicity concentrations, and water levels in soil. Biotic components refer to the living factors in an environment and include the producers, consumers and decomposers. Producers are the organisms that convert inorganic material to organic substances. Plants are an example of a producer. By converting sunlight and carbon dioxide to carbohydrates, they are producing food for the consumers. Consumers and decomposers are completely dependent on producers for nourishment. Since plants are an important producer, it follows that sunlight powers nearly all ecosystems (aquatic and terrestrial).
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Ecology of Organisms
Organisms have adapted and evolved anatomical, physiological, and behavioral characteristics that compensate for variation within the environment. The blending of ecology and evolutionary biology is probably most evident when examining organismal ecology. In this way, the environment shapes the organisms.
Habitat Tolerance
Adaptations that organisms have developed allow them to compensate for minimal temporal and spatial variation withFigure 2: Creatures have learned to thrive in a variety of extreme environments, such as the in their environment. This is accomplished, for example, by regulating body temperature or controlling the rate at which desert climate pictured above. water is transpired. Nevertheless, there are limits to an organism’s ability to compensate for environmental factors. No single species can tolerate all of Earth’s environments. The geographic distribution of a species is thus limited by the physical environment. That is, species distribution is limited by an environment’s abiotic factors.
Although living organisms are found all over the planet, all species have a defined habitat tolerance assigning a viable range of environmental conditions. For example, some plant species can tolerate a broad range of soil variability, while others are confined to a single soil type. If a species has a narrow habitat tolerance because of one or more abiotic factors, then they are limited in their distribution range. Species with a broad range of tolerance are usually distributed widely, whereas those with a narrow range have more restricted distribution. Species can be categorized as specialists or generalists. Specialist species are those that have a limited diet or cannot tolerate a wide range of environmental conditions. Consider a cactus. A cactus has a limited tolerance for soil conditions, water levels and temperatures. Generalists, on the other hand, have a wider range of tolerance for environmental conditions and diet.
For example, omnivores are typically generalists, whereas herbivores tend to be specialists. Interestingly, invasive species are frequently generalists.
Figure 3: Dandelion (Taraxicum officionale)- species like the dandelion
Habitat tolerance along with a species’ geographic range (limited vs. are very common and show no aspects of rarity making them very widespread) and its local population size (large vs. small) determines a common handling a broad range of species’ commonness or rarity. These classifications can be very signifi- tolerances.
cant to industries such as agricultural production and wildlife management.
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Ecology of Organisms
As mentioned above, the environment shapes the organisms. What is the effect of organisms on the environment? Environmental changes are influenced by the organisms that inhabit that environment. In this way, organisms shape the environment. A change in species distribution can modify interactions within the environment. The loss of a native species or the invasion of a non-native species can alter the survival of other organisms within the environment. Therefore, control of invasive species and conservation of endangered species are important to maintain the balance of the entire system.
Pre-Lab Questions
1. Would you expect endangered species to be more frequently generalists or specialists? Explain your answer.
2. How does temperature affect water availability in an ecosystem?
3. Choose a species and describe some adaptations that species developed that allow them to survive in their native habitat.
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Ecology of Organisms
Experiment 1: Effects of pH on Radish Seed Germination
Natural soil pH depends on the parent rock material from which it was formed and processes like climate.
Soil pH is a measure of the acidity or alkalinity of the soil. Acidic soils are considered to have a 5.0 or lower pH value whereas 10.0 or above is considered a strong basic or alkaline soil. The pH of soil affects the solubility of nutrients in soil water and thus it affects the amount of nutrients available for plant uptake. Different nutrients are available under differing pH conditions.
In this experiment we will look at the effect of pH on the germination and growth rate of radish seeds in order to determine the range of pH tolerance for the seed. Acidic or basic water will be used in order to stimulate acidity or alkalinity in soil.
Materials
2 mL 4.5% Acetic Acid (Vinegar), C2H4O2
Soda) Solution, NaHCO3
Permanent Marker
*Paper Towel Sheets (cut to fit into the petri dish)
(3) 5 cm Petri Dishes
*Scissors
3 pH Test Strips
*Sunny Location
Radish Seed Packet
*Water
Ruler
2 mL 15% Saturated Sodium Bicarbonate (Baking *You Must Provide
Procedure
1. Use the permanent marker to label the top of each of the three petri dishes as Acetic Acid, Sodium Bicarbonate, or Water.
2. Carefully cut three small circles from the paper towel sheets. The circles should comfortably fit within the bottom of the petri dish.
3. Place the circles in the dishes, and wet them with approximately 2 mL of each respective solution (acetic acid, sodium bicarbonate, or water).
4. Gently press the reaction pad of three, pH test strips onto the wet paper towels. Record your data in Table 1.
5. Arrange 10 radish seeds on each paper towel in each petri dish. Make sure the seeds have space and are not touching. Then, place the top of the petri dish on the bottom.
6. Create a hypothesis regarding which environment will provide the most hearty radish seed growth. Be sure to indicate why you believe this will be true, and use scientific reasoning. Record your hypothesis in
Post-Lab Question 1.
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Ecology of Organisms
7. Place the petri dishes in a sunny or well-lit, warm place. Be sure to keep the paper towels moist for the length of the experiment with the appropriate solution if any of the towels dry out.
8. Observe the seeds daily for seven days, and record the number of seeds that germinate in Table 1. Note when the seeds crack and roots or shoots emerge). On the seventh day, record the lengths of radish seed sprouts (mm or cm).
Table 1: pH and Radish Seed Germination
Stage/Day Observations
Acetic Acid
Sodium Bicarbonate
Water
Initial pH
1
2
3
4
5
6
7
Post-Lab Questions
1. Record your hypothesis from Step 6 here:
2. Compare and construct a line graph based on the data from Table 1 in the space below. Place the day on the x axis, and the number of seeds germinated on the y axis. Be sure to include a title, label the x and y axes, and provide a legend describing which line corresponds to each plate (e.g., blue = acetic acid, green
= sodium bicarbonate, etc…).
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Ecology of Organisms
3. Was there any noticeable effect on the germination rate of the radish seeds as a result of the pH? Compare and contrast the growth rate for the control with the alkaline and acidic solutions.
4. According to your results would you say that the radish has a broad pH tolerance? Why or why not? Use your data to support your answer.
5. Knowing that acid rain has a pH of 2-3 would you conclude that crop species with a narrow soil pH range are in trouble? Is acid rain a problem for plant species and crops?
6. Research and briefly describe a real world example about how acid rain affect plants. Be sure to demonstrate how pH contributes to the outcome, and proposed solutions (if any). Descriptions should be approximately 2 - 3 paragraphs. Include at least three citations (use APA formatting).
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Lab 28
Ecological Interactions
Ecological Interactions
Learning Objectives
•
Hypothesize how habitat influences niches
•
Explain how substances accumulate in trophic levels through biomagnification
•
Analyze the effects of changes in a food web
Introduction
An ecosystem is composed of communities of living organisms at a particular time. This includes plants and animals, as well as non-living elements such as water, wind, heat, chemical constituents, etc. Each organism within a community has a habitat and a niche. An organism’s habitat is the location in which it physically resides or is adapted to reside. An organism’s niche refers its function or “occupation”. Occupation defines the way in which the organism obtains and sustains all of the elements needed for survival.
Figure 1: Mountain gorillas have a restricted geographic range, a narThe dynamics of an ecosystem and the complexity of its habitats are a row habitat tolerance, and a small result of energy flow, nutrient cycling, and water. Organisms can be prolocal population. It is highly vulneraducers or consumers in terms of energy flow. These distinctions com- ble to extinction.
Trophic Levels
prise the basis for trophic levels. Autotrophs are primary producers; they make their own food typically using water, energy from the environment (e.g., sunlight) and carbon dioxide as their carbon source. In other words, autotrophs do not rely on other organisms for survival; they are self-nourishing. Autotrophs are the critical link between solar radiation or inorganic chemicals and every other planetary consumer. Autotrophs produce all the organic compounds (carbohydrates, fats, proteins) that are critically needed by all other organisms.
Phototrophs use sunlight for their energy source while lithotrophs use inorganic molecules (hydrogen sulfide, sulfur, etc.) for their energy source. Photosynthesis, for example, is used by phototrophs to produce carbohydrates from inorganic molecules as shown in the following equation:
CO2 + H2O + light → CH2O + O2
Heterotrophs are consumers and obtain energy and carbon from organic molecules made by primary producers. Heterotrophs, therefore, rely on other organisms for their nourishment. Heterotrophs are sorted into levels based on what their diet includes. These levels are: primary, secondary and tertiary. Herbivores are primary consumers whereas carnivores and omnivores are secondary consumers. Tertiary consumers are on
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Ecological Interactions top of the food chain and feed on secondary and primary consumers. They are sometime known as apex predators and can be carnivores or omnivores.
Organisms that share the same basic foods are said to be in the same trophic level (feeding level). Beginning with the autotrophs, energy “flows” through the system along a circuit called a food chain. This occurs in the form of carbon-carbon bonds. Carbon-carbon bonds are broken for energy consumption when respiration occurs. For example, corn (a phototroph) produces carbohydrates (starch) which provide humans Figure 2: Heterotrophic species consuming a phototrophic species.
(heterotrophs) with a source of energy and carbon. Consider another scenario in which an herbivore ingests the corn and a predator consumes the herbivore. The herbivore receives its nourishment from the corn and the predator receives its nourishment from the herbivore.
Biomagnification
Biomagnification is the accumulation of a substance as it works its way up the food chain by transfer of the substance from lower trophic level organisms to higher trophic organisms. Biomagnification results in higher or magnified substance concentrations for organisms higher in the food chain. Only about 10% of food calories make it from one trophic level to the next in biotic feeding systems. Energy transfer is in no way 100% efficient. Inefficient energy transfer is what accounts for the classically depicted ecological pyramid, with a wide base representing the producers and a narrow top representing the tertiary consumers.
Figure 3: Inefficient vs. Efficient transfer pyramid. There is a big difference in efficiency between direct and indirect consumption. Calories are lost as they move up the food chain, which also contributes to the fact that there are more prey than predators present in a given ecosystem.
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Ecological Interactions
Organisms at lower trophic levels eat small amounts, but higher trophic levels organisms eat many lower trophic level organisms; thus, chemicals accumulate in higher quantities. Biomagnification becomes a concern when the magnified chemicals are harmful to life or the environment. For example, mercury is not degraded or excreted by organisms. It can be absorbed by autotrophs and ingested by consumers accumulating in tissues. At each level, the concentration of mercury increases.
As mentioned above, all life on Earth is directly or indirectly reliant on primary production. Algae are extremely prevalent autotrophs that undergo photosynthesis (mainly algae) utilize aquatic carbon dioxide and water to produce carbohydrates (CH2O). Recall that photosynthesis produces carbohydrates and oxygen.
Predation and Competition
In an ecosystem , populations interact with each other in a variety of ways. The two most common interactions observed are predation and competition. Competition occurs when two or more populations need the same resource: water, food, or space. Competition can occur directly or indirectly. Predation is an interaction in which one population (prey) is a food source for another (predator).
Symbiotic Relationships
Perhaps some of the most interesting animal behaviors are the long term interactions or relationships observed in nature between two or more different species. Relationships between species range from those which favor only one of the species, to those in which both species benefit. Parasitism is an example of a relationship in which one species benefits and the other species suffers. Mutualism, on the other hand, is a relationship in which members of all involved species benefit. The term symbiosis describes the long-term relationships between organisms of different species. Keep in mind that some resources use the term symbiosis to describe only mutualistic relationships while others use it for all long-term interspecies interactions.
Pre-Lab Questions:
1. Explain how some heterotrophs could be described as either a primary consumer or a secondary consumer.
2. Where do you think fungi fit into a food chain? Are they autotrophs or heterotrophs? Explain your answer using scientific reasoning and data.
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Ecological Interactions
Experiment 1: Ecological Interactions
Materials
(1) 10 mL Graduated Cylinder
contain 1000 mL)
(1) 100 mL Graduated Cylinder
10 mL Vegetable Oil
*You Must Provide
*Large Pitcher, Jug, or Glass (must be able to
Procedure
1. Measure out 990 mL of water into your big jug using the 100 mL graduated cylinder. The jug represents the primary producers.
2. Measure 10 mL of oil and add it to the jug with 990 mL.
3. Clean the 10 mL graduated cylinder so there is no oil residue.
4. Let the oil coalesce at the top of the container, the oil is the accumulation agent. It is insoluble because we want it to represent the organism’s inability to break it down.
5. Calculate the volume of water and the volume of oil in your 1000 mL container. Calculate percent concentration of oil. Record in Table 1.
Hint: V (of two or more substances) = (V (of single substance)/ total V of the mixture) x 100
6. Pour 100 mL of the mixture into the 100 mL graduated cylinder and let the mixture settle.
7. Calculate the volume of water and the volume of oil in your 100 mL container. Calculate the percent concentration of oil. Record in Table 1.
8. Now, pour 10 mL of the mixture into the 10 mL graduated cylinder.
9. Calculate the volume of water and the volume of oil in the 10 mL graduated cylinder. Calculate the percent concentration of oil.
Table 1 Volume and Concentration Totals
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Trophic Level
Cylinder
Volume of
H2O
Volume of Oil Total Volume
1st
1000 mL
1000 mL
2nd
100 mL
100 mL
3rd
10 mL
10 mL
Percent Oil
Ecological Interactions
Post-Lab Questions
1. What is the percent concentration of oil in the first, second, and third trophic levels in our food chain?
2. How did the concentration of oil change from one trophic level to the next?
3. How does the change in concentration represent biomagnification?
4. How does this also illustrate the amount of energy transferred from one trophic level to the next?
If you would like to read more about issues surrounding bioaccumulation simply search for case studies that address POP (persistent organic pollutants) and biomagnification in ecosystems.
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Ecological Interactions
Experiment 2: Oil Spills and Aquatic Animals
Understanding ecological interactions extends beyond reviewing the natural interactions between the environmental and animal factors. In fact, many would argue that human interactions with animals can cause an equally great (or even greater) impact on animals or the environment due to the number of species which become endangered or extinct as a result of human behavior. This may transpire through hunting, habitat encroachment, pollution, etc. In this experiment, you will examine the affect of oil and detergent pollutions animal coats.
Materials
(5) 250 mL Beakers
Sponge
3 Chamois Cloth Pieces
Wooden Stir Stick
3 Feathers
*Stopwatch
(1) 100 mL Graduated Cylinder
*Large Bowl
2
2 /3 T. (apx. 40 mL) Liquid Dishwashing Soap
*Paper Towel
Measuring Spoon
*Water
2
2 /3 T. (apx. 40 mL) Vegetable Oil
Permanent Marker
*You Must Provide
2
2 /3 T. Powdered Dishsoap
2 2/3 T. Powdered Laundry Detergent
Procedure
1. Observe the texture of the chamois cloth. Try to use three to four different adjectives to describe the cloth, and record your observations in Table 3.
2. Repeat Step 1 using the feathers instead of the chamois cloth.
3. Use the permanent marker to label the 250 mL beakers as Liquid Dishwashing Detergent, Powdered
Dishsoap, Powdered Laundry Detergent, Water, Oil + Water.
4. Use the measuring spoon to add 2 2/3 T. (approximately 40 mL) amount of liquid dishwashing detergent, 2
2
/3 T. powdered dishsoap, and 2 2/3 T. powdered laundry into their respective beakers. Set the beakers
aside.
Note: Three total measurements in Step 4. One measurement should be allocated into one of the three, labeled beakers.
5. Use the 100 mL graduated cylinder to add 40 mL water to the Water beaker.
6. Add 100 mL water to the Liquid Dishwashing Detergent, Powdered Dishsoap, and Powdered Laundry Detergent beakers. Use the wooden stir stick to gently stir the mixture, being careful not to introduce unnecessary bubbles to the solutions.
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Ecological Interactions
7. Fill the large bowl approximately 50% with room temperature water. This will be your rinse bowl.
8. Add 40 mL oil and 10 mL water to the Water + Oil beaker.
9. Add the three pieces of chamois cloth to the Water beaker. Allow the cloth to soak for one minute. Note any changes to the cloth texture. Record your observations in Table 3.
10. Add the water-saturated pieces to the Water + Oil beaker. Allow it to soak for an additional three minutes. 11. Rinse the pieces in the Rinse beaker and place them on paper towels to air dry. Note any initial changes to the cloth in Table 3.
12. Repeat Steps 9 - 11 with the feathers.
13. Place one piece of chamois cloth and one feather in the liquid dishwashing detergent beaker. Allow to soak for approximately five minutes.
14. Repeat Step 11 placing items into the Powdered Laundry Detergent and the Powdered Dishwashing
Detergent beakers.
15. Use the sponge to gently wash the items in their respective beakers. Then, individually transfer the items to the rinse bowl and continue to rinse the items.
16. Place each item on a paper towel, and allow to air dry.
17. Examine the dried items and observe their final textural status. How do they compare to the initial texture? Record your observations in Table 3.
Table 3: Cloth and Feather Observations
Item Type
Initial Observations
Water-Saturated
Observations
Oil-Soaked
Observations
Final Observations
Chamois
Cloth
Feather
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Ecological Interactions
Post-Lab Questions
1. Which solution cleaned and restored the items most effectively?
2. Research the ingredients included in powdered laundry detergent, powdered dishwashing soap, and liquid dishwashing soap. What difference(s) do you notice, and how do you think this affected the soaps’ ability to restore the items?
3. Identify one marine species with an outer layer which is similar to the chamois cloth, and one which is similar to the feathers.
4. How might aquatic species be affected in a natural environment if a coat or outer shell was compromised by pollution? How might bird species be affected?
5. Identify one additional way that aquatic species may contact water pollution (other than via coat or shell contamination). 6. Aquatic life may be directly affected when oil leaks or spills occur. For example, the Deepwater Horizon oil spill in 2010 (which is considered to be the largest oil spill in petroleum industry history) affected over
8,000 marine species including fish, birds, mammals, and more. Research one species (plant or animal) that can be affected by oil spills.
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Appendix
Good Lab Techniques
Good Lab Techniques
Good Laboratory Practices
Science labs, whether at universities or in your home, are places of adventure and discovery. One of the first things scientists learn is how exciting experiments can be. However, they must also realize science can be dangerous without some instruction on good laboratory practices.
•
Read the protocol thoroughly before starting any new experiment. You should be familiar with the action required every step of the way.
•
Keep all work spaces free from clutter and dirty dishes.
•
Read the labels on all chemicals, and note the chemical safety rating on each container. Read all Material Safety Data Sheets
(MSDS) prior to each experiment. These are provided on www.eScienceLabs.com. An underpad will prevent any spilled liquids from contaminating the surface you work on.
•
Thoroughly rinse labware (test tubes, beakers, etc.) between experiments. To do so, wash with a soap and hot water solution using a bottle brush to scrub. Rinse completely at least four times. Let air dry.
•
Use a new pipette for each chemical dispensed.
•
Wipe up any chemical spills immediately. Check MSDSs for special handling instructions (provided on www.eScienceLabs.com).
A
B
C
Special measuring tools in make experimentation easier and more accurate in the lab. A shows a beaker, B graduated cylinders, and C test tubes in a test tube rack.
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Good Lab Techniques
•
Use test tube caps or stoppers to cover test tubes when shaking or mixing – not your finger!
•
When preparing a solution, refer to a protocol for any specific instructions on preparation. Weigh out the desired amount of chemicals, and transfer to a beaker or graduated cylinder.
Add LESS than the required amount of water. Swirl or stir to dissolve the chemical (you can also pour the solution back and forth between two test tubes), and once dissolved, transfer to a graduated cylinder and add the required amount of liquid to achieve the final volume.
•
A molar (M) solution is one in which one liter (L) of solution contains the number of grams equal to its molecular weight. Ex:
1 M = 110 g CaCl ÷ 1 L
Disposable pipettes aid in accurate measuring of small volumes of liquids. It is important to use a new pipet for each chemical to avoid contamination. (The formula weight of CaCl is 110 g/mol)
•
A percent solution can be prepared by percentage of weight of chemical to 100 mL of solvent (w/
v) , or volume of chemical in 100ml of solvent (v/v).
Ex:
20 g NaCl ÷ 80 mL H2O = 20% w/v NaCl solution
•
Concentrated solutions, such as 10X, or ten times the normal strength, are diluted such that the final concentration of the solution is 1X.
To make a 100 mL solution of 1X TBE from a 10X solution:
10 mL 10X TBE ÷ 90 mL water = 100 mL 1X TBE
•
Always read the MSDS before disposing of a chemical to insure it does not require extra measures. (provided on www.eScienceLabs.com)
•
Don’t pour unused chemical back into the original bottle.
•
Avoid prolonged exposure of chemicals to direct sunlight and extreme temperatures.
•
.Immediately secure the lid of a chemical after use.
•
Prepare a dilution using the following equation: c1v1 = c2v2
Where c1 is the concentration of the original solution, v1 is the volume of the original solution, and c2 and v2 are the corresponding concentration and volume of the final solution. Since you know c1, c2, and v2, you solve or v1 to figure out how much of the original solution is needed to make a certain volume of a diluted concentration.
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Good Lab Techniques
•
If you are ever required to smell a chemical, always waft a gas toward you, as shown in the image above. This means to wave your hand over the chemical towards you. Never directly smell a chemical. Never smell a gas that is toxic or otherwise dangerous.
•
Use only the chemicals needed for the activity.
•
Keep lids closed when a chemical is not being used.
•
When diluting an acid, always pour the acid into the water. Never pour water into an acid.
•
Never return excess chemical back to the original bottle. This can contaminate the chemical supply.
•
Be careful not to interchange lids between different chemical bottles.
•
When pouring a chemical, always hold the lid of the chemical bottle between your fingers. Never lay the lid down on a surface. This can contaminate the chemical supply.
•
When using knives or blades, always cut away from yourself.
•
Wash your hands after each experiment.
431
Credits
Credits
Text/Photography
Chiras, D., (2010). Environmental science. (8th ed.). Sudbury, Massachusetts: Jones and Bartlett Publishers,
LLC.
Cornely, K., Pratt, C. W., (2004). Essential biochemistry. Hoboken, New Jersey: John Wiley & Sons, Inc.
Damjanov, I., McKinnell, R. G., Parchment, R. E., Perantoni, A. O., & Pierce, G. B. (2006).Biological basis of cancer. (2nd ed.). New York, New York: Cambridge University Press.
Evers, C. A., Starr, C., Starr, L., (2010). Biology today and tomorrow. (3rd ed.). Belmont, California: Brooks/
Cole, Cengage Learning.
Lewis, R., (2007). Human genetics concepts and applications. (7th ed.). New York, New York: The McGraw
Hill Companies, Inc.
Johnson, G. B., Raven, P. H., (1989). Biology. (2nd ed.). St. Louis, Missouri: Times Mirror/Mosby College
Publishing.
Marieb, E. N. (2009). Essentials of human anatomy and physiology. (9th ed.). San Francisco, California:
Pearson Education Inc.
Office of Transportation and Air Quality. (2013, 08 01).Renewable and alternative fuels. Retrieved from http:// www.epa.gov/otaq/fuels/alternative-renewablefuels/ Saferstein, R., (2013). Forensic science from the crime scene to the crime lab. (2nd ed.). San Francisco, California: Pearson Education Inc.
Tiedje, J. M., Thompson, D. K., Xu, Y., & Zhou, J. (1993). J. Zhou (Ed.), Biodiversity Prospecting Hoboken,
New Jersey: Wiley-Liss.
US Department of Energy. (2013, 09 25). Clean energy. Retrieved from http://www.epa.gov/cleanenergy/ energy-and-you/affect/coal.html 433
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