TRACING OF BIOSURFACTANT SYNTHESIZING GENES IN BACILLUS SP., BY IN VITRO AND IN SILICO TECHNIQUES USING SRFA GENE AS MARKER

Abstract

Biosurfactant production enhances the establishment of bacteria in its environment. It was confirmed among the food borne pathogens of Bacillus cereus, B. licheniformis and B. subtilis isolated from spoiled dairy products. Molecular weights of the purified DNA from these isolates were determined as > 4000 Kb. Restriction digestion of extracted genomic DNA by EcoRI and HindIII and amplification by genus specific 16S rRNA derived primer confirmed the homology among all. Production of biosurfactant by these bacteria was confirmed by drops collapse test, reduction in surface tension of culture media and emulsification properties.  Purified biosurfactants from these isolates were characterized as surfactin, lichenysin and plipastatin from B. subtilis, B. licheniformis and ­­B. cereus respectively. BLAST analysis of surfactin synthesizing gene srfA from B. subtilis showed 80% similarity with surfactant coding gene of lichenysin in B. licheniformis, 76% similarity with an unknown non-ribosomal peptidyl protein and 73% with bacitracin synthetase in B. cereus. So, the unknown plipastatin coding genes in B. cereus predicted as a non-ribosomal in origin and have antimicrobial properties.