重组酶Cre基因在大肠杆菌中的高效表达及其一步纯化和活性检测  

Recombinase Cre's overexpression in Escherichia coli BL21(DE3) and its one-step purification and activity assay.

作　　者：王文棋[1] 盖颖[1] 陆海[1] 蒋湘宁[1] 

机构地区：[1]北京林业大学生物科学与技术学院

出　　处：《北京林业大学学报》2006年第3期57-60,共4页Journal of Beijing Forestry University

基　　金：国家自然科学基金项目(30271066;30571512);"973"国家重大基础规划项目(TG1999016005)

摘　　要：为了实现DNA重组酶Cre在体外的应用,以pET-30a高效表达载体为基础构建了pTE30-Cre质粒.将此质粒导入大肠杆菌BL21(DE3)中进行IPTG诱导的Cre基因的高效表达.对表达出的Cre重组酶蛋白进行镍离子鳌合法一步纯化,并以带有两个同向loxP位点的质粒为底物,对纯化出的Cre酶进行活性检测.该实验为Cre酶的体外应用奠定了基础.In order to realize the application of Cre recombinase in vitro, plasmid pTE30-Cre was constructed based on a pET-30a vector. This plasmid was transferred into Escherichia coli BL21 （DE3）. The Cre gene was overexpressed and then induced by IPTG. The Cre recombinase protein was easily purified using a Ni^＋ affinity column and its activity was tested in vitro by providing a plasmid which contained two of the same directional loxP sites.

关 键 词：DNA重组酶Cre pET30-Cre高效表达 一步纯化 Cre酶活性检测 

分 类 号：Q559.1[生物学—生物化学]