ERIC-PCR Analysis
Introduction: Escherichia coli (E.coli) is a commensal-pathogenic organism which include a wide range of strains. There are several advanced molecular-genomic technologies that are capable for detecting and identifying different strains of E.coli. But, ERIC-PCR technique is a quick and cost effective method for determining individual strains via demonstration of strain specific fingerprint bands. Therefore, the ERIC-PCR technique was used to determine the isolated strains of E.coli from different animal stool specimens.
Material and Methods: The animal stool samples from hens, sheep and cows were obtained during 1 year. After screening processes, the E.coli bacteria were isolated and cultured using standard microbiological methods. The DNA …show more content…
Among several PCR based tools, the Enterobacterial repetitive intergenic consensus (ERIC) PCR is a sharp and cost effective molecular-genomic finger printing technology for differentiating genomic dissimilarities among different types of strains. Indeed, ERICs are recognized as mobile DNA particles that are in association with miniature inverted transposable elements (MITEs) (4, 8-14).
The ERIC sequences are recognized in a huge number of bacterial genomes including Enterobacteriaceae family members like E.coli. The incomplete palindrome sequences are generally detected within transcribed areas in association with intergenic regions. Moreover, there are different numbers of ERIC sequence copies among bacterial species (15, 16).
The clonal variability in different bacterial species such as E.coli is achieved throughout the application of primers which are homologous to ERIC sequences. The gained patterns which are appeared in the following of PCR reactions are valuable for evaluating the level of relationships (17). For this reason, the major purpose of present survey is to determine the distribution of ERICs within the isolated strains of E.coli as an appropriate and quick molecular-genomic tool.
Methods and …show more content…
The primers of 5'-ATG TAA GCT CCT GGG GAT TCA C-3' (F) and 5'-AAG TAA GTG ACT GGG GTG AGC G-3' (R) were applied. The process was performed in a volume of 25μl including 1μl bacterial DNA (E.coli), 12.5μl mastermix, 3μl forward and reverse primers (1.5μl/picomol per each), and the left volume was filled by 8.5μl PCR grade water. Finally, thermocycler was programmed according to table 1. Simultaneously, negative (PCR grade water) and positive (bacterial DNA of E.coli) controls were used to have an accurate observation for the results (12,
Material and Methods: The animal stool samples from hens, sheep and cows were obtained during 1 year. After screening processes, the E.coli bacteria were isolated and cultured using standard microbiological methods. The DNA …show more content…
Among several PCR based tools, the Enterobacterial repetitive intergenic consensus (ERIC) PCR is a sharp and cost effective molecular-genomic finger printing technology for differentiating genomic dissimilarities among different types of strains. Indeed, ERICs are recognized as mobile DNA particles that are in association with miniature inverted transposable elements (MITEs) (4, 8-14).
The ERIC sequences are recognized in a huge number of bacterial genomes including Enterobacteriaceae family members like E.coli. The incomplete palindrome sequences are generally detected within transcribed areas in association with intergenic regions. Moreover, there are different numbers of ERIC sequence copies among bacterial species (15, 16).
The clonal variability in different bacterial species such as E.coli is achieved throughout the application of primers which are homologous to ERIC sequences. The gained patterns which are appeared in the following of PCR reactions are valuable for evaluating the level of relationships (17). For this reason, the major purpose of present survey is to determine the distribution of ERICs within the isolated strains of E.coli as an appropriate and quick molecular-genomic tool.
Methods and …show more content…
The primers of 5'-ATG TAA GCT CCT GGG GAT TCA C-3' (F) and 5'-AAG TAA GTG ACT GGG GTG AGC G-3' (R) were applied. The process was performed in a volume of 25μl including 1μl bacterial DNA (E.coli), 12.5μl mastermix, 3μl forward and reverse primers (1.5μl/picomol per each), and the left volume was filled by 8.5μl PCR grade water. Finally, thermocycler was programmed according to table 1. Simultaneously, negative (PCR grade water) and positive (bacterial DNA of E.coli) controls were used to have an accurate observation for the results (12,