Construction Of Engineering Bacillus Cereus Strain And Expressing α-amylase Gene

Bacillus cereus can produceα-amylase which has important industrial value and can endure high temperature. The enzymatic characteristic of twoα-amylase from B.cereus (905 and 904)were studied. The results showed that: the activity stability temperature of bothα-amylase is 90℃-100℃. Their thermal stability and enzymatic activity do not depend on Ca2+. But their protein molecular weights are obviously different. The structural gene of amy905 and amy904 were successfully cloned by PCR and were successfully expressed in E. coli. The experiments showed that, the gene of amy904 is 1362bp and the protein molecular weight is about 55 KD; the gene of amy905 is 1761bp and the protein molecular weight is about 68 KD.The gene of amy904 was inserted into the E.coli-Bacillus shuttle vector pHP14-promp to construct the vector of pHP14-amy904. The new vector was transformed into B.cereus905 and was expressed in B.cereus905 by electroporation. The stability on heredity has been evaluated in this experiment; the result showed that the stability is 93%. This result implied that the strain of B.cereus905-amy904 strain could satisfy to the study during the whole period of crop. The growth speed was similar between wild and B.cereus905-amy904 stains by inspecting growth curves, which suggested that the transformation of pHP14-amy904 did not affect the metabolism of B. subtilis greatly.