Arabinoe Operon Promoter
Page 1
Analysis of transcriptional regulation of the arabinose operon promoter using expression from a green fluorescent protein encoding reporter gene
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KIRA FERNANDEZ
, CLAIRE MARKEY
*
, JESSE PIERATTI
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Department of Biological Sciences
, and Department of Nutrition and Dietetics*, Messiah
College, Mechanicsburg, PA 17055
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ABSTRACT
This study investigates gene regulation and how environmental arabinose and/or glucose can interact with genotype to influence phenotypic expression of the Green Fluorescent
Protein (GFP) gene inserted in the plasmid under the control of a promoter called the pBAD promoter. Bacterial cells are a common choice for in vivo replication of DNA of interest, and in …show more content…
The more the bacteria used the glucose, the less of it was around to repress the operon, which is why its fluorescence strength grew over time. If the study was to be continued past the 96 hour mark, all of the plates would have eventually fluoresced as they used up their glucose resources and began activating the arabinose operon (4).
Bacterial Transformation
The plate containing only arabinose and lysogeny broth (LB) was overgrown by bacteria, creating a lawn of colonies. This is because nothing was added to separate a bacterium that took up a plasmid from a bacterium that had not. All of the bacteria spread on the plate were able to use the lysogeny broth and arabinose to grow and form visible colonies. The 10 μL sample produced only one nonfluorescent colony. The total of fluorescent cells in the 100μL and 100μL
(5x) samples were around ⅕ of the total cells grown on the plate. In these three samples, ampicillin was added to the plates to kill any bacteria that did not take up a plasmid. The bacteria that did take up a plasmid became ampicillin resistant from a selectable marker gene that was placed into the plasmid for this purpose (4).
Not all of the bacteria that took up a plasmid and survived the ampicillin glowed. This
Analysis of transcriptional regulation of the arabinose operon promoter using expression from a green fluorescent protein encoding reporter gene
+
+
KIRA FERNANDEZ
, CLAIRE MARKEY
*
, JESSE PIERATTI
+
Department of Biological Sciences
, and Department of Nutrition and Dietetics*, Messiah
College, Mechanicsburg, PA 17055
Page 2
ABSTRACT
This study investigates gene regulation and how environmental arabinose and/or glucose can interact with genotype to influence phenotypic expression of the Green Fluorescent
Protein (GFP) gene inserted in the plasmid under the control of a promoter called the pBAD promoter. Bacterial cells are a common choice for in vivo replication of DNA of interest, and in …show more content…
The more the bacteria used the glucose, the less of it was around to repress the operon, which is why its fluorescence strength grew over time. If the study was to be continued past the 96 hour mark, all of the plates would have eventually fluoresced as they used up their glucose resources and began activating the arabinose operon (4).
Bacterial Transformation
The plate containing only arabinose and lysogeny broth (LB) was overgrown by bacteria, creating a lawn of colonies. This is because nothing was added to separate a bacterium that took up a plasmid from a bacterium that had not. All of the bacteria spread on the plate were able to use the lysogeny broth and arabinose to grow and form visible colonies. The 10 μL sample produced only one nonfluorescent colony. The total of fluorescent cells in the 100μL and 100μL
(5x) samples were around ⅕ of the total cells grown on the plate. In these three samples, ampicillin was added to the plates to kill any bacteria that did not take up a plasmid. The bacteria that did take up a plasmid became ampicillin resistant from a selectable marker gene that was placed into the plasmid for this purpose (4).
Not all of the bacteria that took up a plasmid and survived the ampicillin glowed. This