MOLECULAR IDENTIFICATION AND CHARACTERIZATION OF POLY-β-HYDROXYBUTYRATE (PHB) PRODUCING BACTERIA ISOLATED FROM CONTAMINATED SOILSKIRAN NEHRA, NANCY CHHABRA, PARVEEN K. SIDHU, PRIYANKA LATHWAL AND J.S. RANA
Polyhydroxybutyrate (PHB) producing bacteria are an economically important source for the production of PHB, a compound having the potential to replace non-degradable conventional plastics with biodegradable plastics. The present study deals with the molecular characterization of two PHB positive bacterial isolates, IBB and ITG, isolated from industrial effluent discharge sites. PHB extraction carried out using sodium hypochlorite digestion method, exhibited a good PHB yield amounting to 82.53 mg/mL and 63.35 mg/mL for IBB and ITG isolates, respectively. The percent PHB accumulation was observed to be 68.38% and 62.76 %, respectively, for IBB and ITG isolates. For identification, the isolates were initially subjected to various standard biochemical tests which were then followed by their molecular characterization. Based upon 16S rRNA gene amplification and phylogenetic relationship studies, the two isolates were identified as Bacillus anthracis (IBB) and Bacillus subtilis (ITG). Further, polymerase chain reaction (PCR) protocol was designed for detection of the gene encoding polyhydroxybutyrate synthase enzyme. PHB synthase gene specific primers were designed on the basis of published sequences of the gene, and these were found to exhibit amplification of a 1089 bp fragment (representing phaC gene) from the genomic DNA of the isolates studied; thus, confirming the presence of PHB producing genes in the two isolates.
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