文档介绍：Cloning and expression of a thermostable pullulanase
gene from Thermotoga maritima MSB8 in Bacillus
subtilis WB600
Zhe Su . Fu-Ping Lu﹡. Qiang Gao . Xiao-Guang Liu . Bin-Zhe Wang . Tao Niu
Key Laboratory of Industrial Microbiology, Ministry of Education,
College of Biotechnology, Tianjin University of Science and Technology
Tianjin 300457, P. R. China
marilyn66@, ******@tust.
Abstract—The pulA gene enconding a thermostable pullulanase
of 844 amino acids was cloned by PCR amplification from the II. MATERIALS AND METHODS
genomic DNA of Thermotoga maritima MSB8. We essfully
expressed the pulA gene in a protease-deficient strain Bacillus A. Strains and Plasmids
subtilis WB600, and the expressed pullulanase was secreted into Strains Escherichia coli JM109, B. subtilis WB600 and the
the culture broth. Preliminary results showed that the plasmid pWB980 were from our lab collections. Vector
pullulanase exhibited optimal activity at 90°C, and was stable at pMD19-T Simple was purchased from TaKaRa Biotechnology
the temperature range from 30 to 80°C for 30 min. The stable pH (Dalian) Co., Ltd.
range for this pullulanase was from to , with an optimum
at pH .
B. Growth Conditions
Keywords-Pullulanase; Thermotoga maritima MSB8; Bacillus E. coli JM109 and B. subtilis WB600 were cultivated
subtilis WB600; pWB980; Enzymatic proper