Zhang Jiayu, Wu Dan, Li Zhaofeng, Chen Sheng, Chen Jian, Wu Jing
Key Laboratory of Industrial Biotechnology, Ministry of Education, School of Biotechnology, Jiangnan University, Wuxi 214122, China.
Sheng Wu Gong Cheng Xue Bao. 2009 Dec;25(12):1948-54.
The cgt gene was isolated from Paenibacillus macerans by PCR amplification and was inserted into vectors of pPIC9K and pMAS. The recombinant vectors were transformed to Pichia pastoris KM71 and Bacillus subtilis WB600, respectively. The results showed that alpha-CGTase activity in the culture media of recombinant P pastoris was only 0.2 U/mL, while it was 1.9 U/mL in recombinant B. subtilis. In addition, we optimized the culture conditions of the recombinant B. subtilis strain. After cultivation at 37 degrees C for 24 h with shake flask, the CGTase forming activity in culture media reached to 4.5 U/mL (hydrolysis activity was 3200 IU/mL), which is 9.8-fold to that of the original strain P. macerans.
通过PCR扩增从浸麻芽孢杆菌中分离出cgt基因，并将其插入pPIC9K和pMAS载体中。将重组载体分别转化到毕赤酵母KM71和枯草芽孢杆菌WB600中。结果表明，重组毕赤酵母培养基中的α-环糊精葡萄糖基转移酶（α-CGTase）活性仅为0.2 U/mL，而重组枯草芽孢杆菌中的活性为1.9 U/mL。此外，我们优化了重组枯草芽孢杆菌菌株的培养条件。在摇瓶中于37℃培养24小时后，培养基中的CGTase形成活性达到4.5 U/mL（水解活性为3200 IU/mL），是原始菌株浸麻芽孢杆菌的9.8倍。
