人era基因的克隆测序及表达

Cloning, sequencing and expression of cDNA gene of human era

目的 克隆人的 era基因 (简称 Hera)并利用大肠杆菌进行表达 .方法 PCR扩增 Hera基因 ,测序正确后克隆入大肠杆菌融合表达载体 p GEX- 4T3,受控于 Ptac启动子 ,重组质粒 p GEX- Hera以大肠杆菌 DH5α为宿主菌 ,用 IPTG进行诱导表达 .结果 克隆了 Hera基因 ,测序正确 ;含重组质粒p GEX- Hera的菌体诱导后在 SDS- PAGE上出现一条新生蛋白带 ,相对分子质量为 6 5 ku,占菌体总蛋白的 2 3% .结论 成功扩增、克隆 Hera基因 。

AIM To amplify and to clone the human era gene and to express its recombinant protein in E.coli. METHODS After the human era gene was amplified by PCR and identified by sequencing, it was inserted into expression vector pGEX4T3 in which exogenous gene was controlled by ptac promoters. The recombinant plasmid pGEXHera was transformed into DH5α and induced with IPTG chemically. RESULTS The human and sequenced correctly. When the engineered bacteria were induced, an anticipated 65 ku protein band appeared on SDSnted to 23% of total bacterial protein. CONCLUSION The human era gene has been successfully subcloned and efficiently expressed in E.coli.