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Title

THE STUDY OF BIODIVERSITY OF PHENOL-DEGRADING BACTERIA BY REP-PCR

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Abstract

 RECENT PROGRESS IN MOLECULAR MICROBIAL ECOLOGY HAS REVEALED THAT TRADITIONAL CULTURING METHODS FAIL TO REPRESENT THE SCOPE OF MICROBIAL DIVERSITY IN NATURE, SINCE ONLY A SMALL PROPORTION OF VIABLE MICROORGANISMS IN A SAMPLE ARE RECOVERED BY CULTURING TECHNIQUES. TO DEVELOP METHODS TO INVESTIGATE THE FULL EXTENT OF MICROBIAL DIVERSITY, WE USED METAGENOMIC STUDIES FOR PHENOL DEGRADING BACTERIA IN A PETROLEUM CONTAMINATED SOIL. METAGENOMICS IS “AN APPLICATION OF MODERN GENOMICS TECHNIQUES TO THE STUDY OF COMMUNITIES OF MICROBIAL ORGANISMS DIRECTLY IN THEIR NATURAL ENVIRONMENTS, BYPASSING THE NEED FOR ISOLATION AND LAB CULTIVATION OF INDIVIDUAL SPECIES”.THEREFORE, IN THIS METAGENOMIC STUDY, AND TO DEMONSTRATETHE DIVERSITY OF ISOLATED BACTERIA, GENOMIC FINGERPRINTING WAS INVESTIGATED BYREPETITIVE EXTRAGENIC PALINDROMIC SEQUENCE PCR (REP-PCR).HOWEVER, BACTERIA WERE ISOLATED FROM CONTAMINATED SOIL BY PLATING AFTER ENRICHMENT IN BATCH CULTURES. THEN, TOTAL BACTERIAL DNA WAS EXTRACTED BY SET-BUFFER METHOD AND REP-PCR PERFORMED WITH PRIMERS REP1R-I AND REP2-I. DIFFERENT PATTERNS WERE OBTAINED BY SEPARATION OF PCR PRODUCTS BY AGAROSE GEL ELECTROPHORESIS. HOWEVER, THE REP-PCR ANALYSIS WAS REPEATED SEVERAL TIMES TO DETERMINE THE REPRODUCIBILITYOF THE METHOD. THE BACTERIAL CELLS ISOLATED WERE CLASSIFIED INTO 18 DISTINCT GROUPS BY A REPETITIVE EXTRAGENIC PALINDROMIC SEQUENCE PCR ANALYSIS. IT IS ESTABLISHED THAT MOLECULAR APPROACHES HAVEEXPANDED OUR KNOWLEDGE OF THE DIVERSITY AND DISTRIBUTIONOF MICROBIAL POPULATIONS IN THE ENVIRONMENT. ON THE OTHER HAND, THE REP PCR COULD BECOMEA POWERFUL TOOL FOR THE MOLECULAR GENETIC ANALYSIS OF BACTERIAAND FOR BACTERIAL TAXONOMY, SINCE IT ALLOWS THE FINGERPRINTINGOF INDIVIDUAL GENERA, SPECIES, AND STRAINS AND COULD HELPDETERMINE PHYLOGENETIC RELATIONSHIPS. THE SAME IS EXPECTED IN METAGENOMICS.

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