Isolation, Identification and Optimization of Culture Conditions of Bacillus sp. Strain PM1 for Alkalo-thermostable Amylase Production.

ABSTRACT

Aims: To isolate and optimize the culture conditions for thermo stable and alkaline amylase production from bacteria. Study Design: Optimization of different physiological and nutritional parameters for amylase production and kinetic studies of amylase. Place and Duration of Study Soil Samples Herbal garden of Amity University Haryana, Gurgaon (Manesar), India, between April 2012 and September 2012. Methodology: Amylolytic isolates were selected by flooding the nutrient agar plates containing 2% starch with Lugol solution. Isolates were selected on the basis of higher ratio of clear zone to colony size and grown in nutrient broth containing 2% starch. The level of amylase was detected in the culture filtrate. The selected isolate showing maximum amylase production was identified on the basis of 16S rDNA amplification. Results: An Alkalo-thermostable amylase producing bacterial isolate from soil was identified as Bacillus sp. strain PM1 on the basis of 16S rRNA. It yielded 3.5 U/ml of amylase in medium containing (%) starch 2.0, beef extract 0.5, NaCl 0.5 at 50ºC, pH 7.0 at 180 rpm after 72 h. The optimum pH and temperature for amylase activity was 8.0 and 50°C, respectively. The enzyme exhibited 67% activity after 60 minute incubation at 50ºC. At pH 8.0, the enzyme retained 78% activity after 4 h. Conclusion: The properties of the isolated enzyme are adequate for its use in starch processing and baking industry.