Cloning and characterization of α-amylase from Bacillus circulans ATCC 61

Amylases are one of the most important enzymes and have a great significance in the present industry of biotechnology. Enzymes obtained from microorganisms are widely used in the industrial field. In the present work, Bacillus circulans ATCC 61 for 𝛼-amylase production has been used. Because well known as a producer of 𝛼-amylase and its genome structures has been known very well, it was used for cloning for this purpose, the genomic DNA of Bacillus circulans ATCC 61 was isolated and the gene encoding α-amylase enzyme was amplified with specific primer by PCR consist of 1400 bp long. The specific primer was designed while detecting the gene belonging to Bacillus α-amylase from gene library. The gel-purified PCR product was inserted into the pGEM®-T Easy vector3015 bp long, which linearized vectors with a single 3´-terminal thymidine at both ends. The T-overhangs at the insertion site greatly improve the efficiency of ligation of A-tailing on the PCR products. Taq DNA polymerase independently generates single deoxyadenosine at both ends of PCR product. Bioinformatics analysis of the cloned gene showed that there is 100% similarity between this gene and amylase gene of Bacillus sp. These results display that the target gene was cloned successfully. Finally for the gene expression, the vector was transferred to JM109 high efficiency E. coli competent cells. This study is the first record in term of the cloning α- amylase enzyme from Bacillus circulans ATCC 61 in the literature.