cre基因在大肠杆菌中的表达及表达蛋白活性的检测

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-29a，在大肠杆菌BL21（DE3）得到了高效表达，采用DEAE－52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。

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

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.