Analysis, Optimization and Molecular Characterization of PHB Positive Bacteria Isolated from Agricultural Soil Sample

Abstract

Polyhydroxybutyrates are the widely studied biopolymer because of its biodegradability and non-toxic effect in the environment. This work focused on the isolation, screening, characterization and optimization of the PHB producing bacteria from agricultural soil. About eighty percentage of the isolated bacteria were identified as PHB producing bacteria after Sudan Black Blue staining. Among them three colonies were randomly selected and their morphologically and biochemically characterized. The biopolymer was extracted using sodium hypochlorite method. Potent PHB producer (WJSP1) (13.5µg/ml) was identified by crotonic acid assay. The functional groups of the produced PHB were identified by Fourier Transform Infra-Red spectroscopy (FTIR). The presence of functional groups like carbon group, C=O ester group, CH group, carbonyl group, carboxyl group which was corelated with the peak values1454.33cm^-1, 1724.36 cm^-1, 1274.95 cm^-1, 1049.28 cm^-1, 1280-1053 cm^-1 indicates the presence of PHB. The potent producer (WJSP1) was optimized under different conditions to achieve the best condition for the production of PHB. The maximum production was obtained at the temperature - 37˚C, the carbon source - glucose, pH - 7 and at a time period of 72 hours. 16SrRNA sequencing was performed and the organism was identified as Bacillus cereus from the BLST (Basic Local Alignment Search Tool). A phylogenetic tree was constructed and the sequence was submitted in GENBANK (Accession No: OR192860). The isolated bacteria Bacillus cereus is an efficient producer of PHB under laboratory condition in presence of glucose as a chief carbon source. Large scale production of PHB can be done in large scale fermenters in industries. Large scale production of PHB will help the environment to get rid of the petroleum-based bioplastics.