β-甘露聚糖酶基因高效表达体系的构建  

The Construction of High-effective Expression System of β-Mannanase Gene

作　　者：李斌[1] 刘正初[1] 王溪森[1] 石岩[1] 

机构地区：[1]中国农业科学院麻类研究所,长沙410205

出　　处：《湖北农业科学》2009年第8期1807-1810,共4页Hubei Agricultural Sciences

基　　金：国家"863"计划项目(2006AA02Z249)

摘　　要：为构建β-甘露聚糖酶基因高效表达体系,采用重叠区扩增基因拼接法将苏云金芽孢杆菌(Bacillus thuringiensis)CTC S-层蛋白启动子和欧文氏杆菌(Erwinia carotovora)CXJZ95-198 β-甘露聚糖酶基因完整开放阅读框定向连接,得到基因拼接体slp-man。将slp-man克隆到高效表达载体pET-28a+上,并转化到大肠杆菌(Escherichia coli)JM109中。结果表明,β-甘露聚糖酶基因高效表达体系构建成功,重组菌发酵11h酶活达到671.3U·mL-1,是欧文氏杆菌CXJZ95-198的1.48倍。To clone and express the gene recombinant sip-man in Escherict, ia coli JMI09, the strong promoter of Bacillus thuringiensis CTC surface layer protein and the β-mannanase gene ORF of Erwinia carotovora CXJZ95-198 were ligated via gene splicing by overlap extension （SOEing）. The obtained directional gene splice of slp-man was then inserted into the high-effective expression vector pET-28a＋. The recombinant plasmid was transformed into Escherichia coli JM109. The result showed that a high-effective expression system of β-mannanase was constructed in Escherichia coli JM109. The β-mannanase activity of the recombinant strain was up to 671.3U.mL^-1 after llh fermentation, which was as 1.48 times as that in E. carotovora CXJZ95-198.

关 键 词：Β-甘露聚糖酶基因 欧文氏杆菌 重叠区扩增基因拼接法 

分 类 号：S188[农业科学—农业基础科学] Q789[生物学—分子生物学]