重组苏云金芽孢杆菌丙酮酸脱氢酶克隆及表达条件的优化  

The Cloning and Expression Optimization of Recombined Bacillus Thuringiensis Pyruvate Dehydrogenase

作　　者：杨迪[1] 李艳[1] 张喜红[1] 袁均林[1] 贺红武[1] 

机构地区：[1]华中师范大学生命科学学院环境科学实验室,湖北武汉430079

出　　处：《化学与生物工程》2009年第4期39-42,共4页Chemistry & Bioengineering

基　　金：国家科技部支撑计划资助项目(2006BAJ02A10);国家重点基础研究发展计划(973计划)资助项目(2003CB114406)

摘　　要：根据Bcl4579丙酮酸脱氢酶基因序列设计引物，从苏云金芽孢杆菌BMB171菌株总DNA中扩增得到相应的丙酮酸脱氢酶基因DNA片段。将DNA片段装载到大肠杆菌构建pET—E1表达系统，再通过优化重组菌的表达条件获得有生物活性的丙酮酸脱氢酶。结果表明，pET—E1表达系统构建获得成功；优化的表达条件如下：培养基为TB＋M9（体积比1：1）、起始菌体密度OD600为4～5．5、诱导剂IPTG浓度为0．04mmol·L^-1。With the primers designed according to the pdhAB gene sequence of Bc14579, the corresponding pyruvate dehydrogenase gene DNA fragments were cloned by amplication of total DNA of Bt strain BMB171 and were assembled in E. coli to construct pET-E1 expression system. Afterwards, pyruvate dehydrogenase with biological activity could be obtained by optimization of expression conditions for recombinant bacterium. The re- sults showed that, pET-E1 expression system had been constructed successfully, and the optimum expression conditions were as follows.the culture medium was TB＋M9 with volume ratio of 1 ： 1,the initial cell density （OD600） was 445.5 and the concentration of inducer IPTG was 0.04 mmol · L^-1.

关 键 词：丙酮酸脱氢酶 表达 克隆 苏云金芽孢杆菌 

分 类 号：TQ920.1[轻工技术与工程—发酵工程] Q786[生物学—分子生物学]