过氧化氢酶基因在大肠杆菌中的克隆表达及发酵优化  

Cloning,expression and fermentation optimization of a catalase gene in E.coli

作　　者：周丽萍[1] 张东旭[1] 李江华[1] 堵国成[1,2] 陈坚[1,2] 

机构地区：[1]江南大学工业生物技术教育部重点实验室,江苏无锡214122 [2]江南大学食品科学与技术国家重点实验室,江苏无锡214122

出　　处：《工业微生物》2011年第3期66-70,共5页Industrial Microbiology

基　　金：国家自然科学基金重点项目(20836003);973项目(2007CB714306);教育部新世纪优秀人才支持计划(NCET-07-0378)

摘　　要：利用PCR扩增技术得到枯草芽孢杆菌(Bacillus subtilis)过氧化氢酶基因katA,将该基因与表达载体pET-20b(+)连接构建重组质粒,经测序验证后,在大肠杆菌JM109中进行表达得到重组大肠杆菌基因工程菌E.coli BL21(DE3)(pET-20b(+)-katA)。SDS-PAGE电泳结果显示出明显的特异性表达条带,大小与过氧化氢酶吻合。摇瓶实验获得重组菌的最佳碳、氮源成分:5 g/L甘油,40 g/L安琪酵母粉,产酶水平最高达到20 000 U/mL。The catalase gene katA from Bacillus subtilis WSHDZ-01 was amplified by PCR. It was then inserted into the expression vector pET-20b（ ＋ ） to create the recombinant plasmid pET-20b（ ＋ ）-katA and the modified vector was transformed into E. coli JM109 for expression. The size of the expressed specific band by SDS-PAGE electrophoresis corresponded wetl to that of the catalase gene katA. For batch fermentation in flasks, the medium composition regarding the carbon and nitrogen was optimized and the results indicated that the maximal enzyme activity of 20 000 U/mL was obtained with 5 g/L glycerol and 40 g/L yeast extract, respectively.

关 键 词：过氧化氢酶 枯草芽孢杆菌 质粒载体 发酵优化 

分 类 号：Q78[生物学—分子生物学] TQ920[轻工技术与工程—发酵工程]