Cloning and expression of an amylase gene from Bacillus sp. isolated from an agricultural field in West Bengal, India

Abstract

The use of fertilizer is pivotal to ensure soil fertility. Green fertilizers have many advantages and bio-molecules generated in a bacterial cell that can be used as green fertilizer. To produce bio-molecules from a bacterial cell, a bacterium was isolated that uses natural starch as sole carbon source. The organism was grown in presence of various natural starchy materials. The organism was partially identified as Bacillus sp. The optimum conditions for growth of the strain were: temperature of 37.0°C and pH 7.0. However, it grows well even above 55.0°C. The optimum conditions for degradation of various starchy substrates using extra-cellular amylase of the strain were studied. The optimum pH was 4.0 for soluble starch whereas it was pH 5.0 for other substrates and temperature was 17.0°C for soluble starch and 37.0°C for others. Prolongation of time of incubation, velocity of enzyme reaction, and optimum concentration of various metal ions for amylase activity was also studied using optimum assay conditions. Molecular cloning and expression of amy gene of the strain was done in Escherichia coli DH5α. The host cell harboring the recombinant plasmid was grown in the presence of potato starch and incubated for many days to obtained cellular lysate and the liquid biomass might be used as bio-fertilizer.

 Keywords: Amylolytic bacteria, amylase, bio-molecules, bio-fertilizer, single cell fertilizer.

African Journal of Biotechnolo, Vol 13(31) 3281-3291