Screening streptomycin resistant mutations from gamma ray irradiated Bacillus subtilis B5 for selection of potential mutants with high production of protease

Bacillus subtilis B5 - a high protease-producing bacterium, was selected from various domestic laboratories. In this study, the effect of gamma irradiation on survival rate and mutant frequency of antibiotic resistance of Bacillus subtilis B5 were determined. The results showed that the survival rate of bacteria decreased with increasing dose of gamma rays and the mutant frequency of Bacillus subtilis increased with irradiation at the higher dose. The highest mutant frequency was 1.61x10^-3 at 1000 Gy (1 mutation for 621 CFU for 20 µg/ml of streptomycin), while it was only 3.09x10^-6 (1 mutation for 0.323x10⁶ CFU for 20 µg/ml of streptomycin) at 100 Gy. 25 colonies, which can produce protease more than the original strain, were initially screened from 361 colonies that are resistant to gamma radiation and 20 µg/ml of streptomycin. Sequencing showed that 3/25 potential colonies were detected rpoB mutated gene (double mutations occur in two of those and single mutation occurs in one remain). (author)