cre基因在大肠杆菌中的表达及表达蛋白活性的检测  

Expression of cre Gene in Escherichia coli and Bioassay Its Expression Product

作　　者：王立霞[1] 张珠强 胡晓倩[2] 胡鸢雷[1] 高音[1] 林忠平[1] 

机构地区：[1]北京大学蛋白质工程及植物基因工程国家重点实验室 [2]北京大学生化及分子生物学系,北京100871

出　　处：《生物工程学报》2002年第4期497-500,共4页Chinese Journal of Biotechnology

基　　金：国家高技术 86 3计划资助项目 (No.0 10 0 4 0 3 0 1)~~

摘　　要：Cre重组酶来自噬菌体P1,可以识别特异的loxP位点的DNA序列 ,并进行专一性的剪切和拼接。利用PCR技术将cre基因克隆至原核表达载体pET 2 9a ,在大肠杆菌BL2 1(DE3)得到了高效表达。采用DEAE 5 2柱层析的方法对表达蛋白进行了纯化。体外生物学活性检测表明 ,表达蛋白对含有同向loxP位点的质粒有切割活性。The Cre recombinase from bacteriophage P1 can recognize specific DNA sequences, cleave DNA at specific target sites, and then ligate it to the cleaved DNA of a second site. In this study, cre gene was cloned into the pGEM-T Easy vector via PCR procedure. Then the cre gene was inserted into an expression vector pET-29a and expressed in E.coli BL21(DE3). A 38kD soluble protein was expressed and named CRE. CRE was purified by DEAE-52 chromatography. Bioassay of the partially purified product showed that CRE can cleave the plasmid pGLGFP which contains two loxP sites with the same direction.

关 键 词：CRE基因 大肠杆菌 表达蛋白活性 检测 CRE重组酶 基因表达 剪切活性 

分 类 号：Q786[生物学—分子生物学]