Mutant strain improvement from bacillus subtilis E6-5 by physi̇cal mutagenesis (ultraviyole, UV) and chemical mutagenesis (Ethidium bromide, EtBr) for enhanced production of protease and optimization of growth medium

In this study, the parent strain Bacillus subtilis E6-5 was exposed to UV and EtBr treatment by classical mutation procedure, and total 200 mutants were obtained. Among of these mutants, a mutant strain was selected based on its ability to produce highest zone formation (25 mm), and strain was named as Bacillus subtilis ATA38. Obtained mutant was tested for enzyme production capacity in the liquid media, the mutant ATA38 showed 18.2 fold increase in protease production over the parent strain B.subtilis E6-5. It was found that mutant strain produced protease on maximum level at 24 th hour. In addition, the effects of nutrional and physical factors on enzyme production were examined. For this purpose, the effects of major medium ingredients (different carbon sources, nitrogen sources and metal ions) on the production of the enzyme were used. The best carbon source was found to be wheat starch (525 IU/mL). Among the organic nitrogen sources, the highest enzyme production was obtained in the presence of meat extract (850 IU/mL),KCl+CaCl2 (528 IU/mL) was detected as effective as metal ion. In the physical parameters, the best results was obtained at 37°C, pH 6.0, 200 rpm asagitation rate, 1% as inoculum size and 18 hours as inoculum age. A new medium was obtainedby optimizing the incubation conditions (nutrional and physical factors) of protease production from ATA38 mutant. In this medium, enzyme yield was enhanced 171% compared to basal medium. The production of protease by ATA38 was inreased 18.2 fold compared to the parental strain. The mutant strain produced may have a prominent potential for protease production on an industrial scale. Key words: Bacillus, Protease, Mutagenesis, UV, EtBr, Nutritional Factors, Physical Factors