Cloning of the thermostable alpha-amylase gene from Bacillus sp. 1-15 | Клонирование гена термостабильной альфа-амилазы Bacillus SP. 1-15
2012
Kachan, A.V. | Evtushenkov, A.N., Belarus State Univ., Minsk (Belarus)
Enzymes, realizing hydrolysis of starch, are important substances, used in modern biotechnology. Alpha-amylases appear the compounds of high commercial value, corresponding several requirements, such as high temperature stability along with high and low pH activity. Alpha-amylase enzymes are isolated out of different organisms; but for industrial necessities are usually used bacterial alpha-amylases, produced by Bacillus and Aspergillus. The main goal of the research is cloning of the gene of thermostable amylase from cryptogamic bacteria into the cells of Escherichia coli and determination of its' nucleotide and amino acid sequences. Morphological and biochemical characteristics of a new native isolate Bacillus sp. 1-15 is carried out in the study. The isolated strain produces thermostable amyolytic enzymes. The gene encoding the amylase from bacillus sp. 1-15 strain is isolated by PCR and cloned in Escherichia coli BL21 (DE3) cells in pET24b-amyl-15 expression vector. Production of the active recombinant enzyme is confirmed. The nucleotide sequence of the Bacillus sp. 1-15 amylase gene is determined. The coding sequence of the gene consist of 1536 base pairs is highly homologous with genes of alpha-amylases from B. lichenifoemis strains.
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