人Era蛋白在大肠杆菌中的高效表达及其纯化

目的 在大肠杆菌中对人era基因（简称hera)进行表达、纯化。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pRSET-C，重组质粒pRSETC-hera以大肠杆菌BL21（DE3）为宿主菌，用IPTG进行诱导表达。然后利用亲和层析的方法对表达蛋白进行纯化。结果 克隆了hera基因，测序正确；利用pRSET-C载体在大肠杆菌中融合表达了全长hera-cDNA基因，表达量占细菌总蛋白的59．6％。融合蛋白在菌体内主要以包涵体形成存在，在变性条件下用Ni-NTA亲和色谱柱纯化此融合蛋白，其纯度达到了98．0％。结论 利用基因重组技术在大肠杆菌中高表达了hera基因。并对融合蛋白进行了初步纯化。

High expression of human Era protein and its purification

AIM To express human Era protein in E.coli and purify it preliminarily. METHODS After being amplified by PCR and identified by sequencing the human era gene was inserted into expression vector pRSET C. The recombinant plasmid pRSETC h era was transformed into BL21(DE3) and induced with IPTG. The expressed protein was purified by affinity chromotography. RESULTS The human era gene was amplified and the sequence proved to be correct. The h era cDNA gene had been cloned into the vector and expressed in E.coli , and the expressed protein took 59.6% of the total bacterial protein. The fused protein existed in the pattern of inclusion body, and it was purified on denatured condition by using Ni NTA affinity chromotography column. CONCLUSION The h era gene could be expressed with high efficiency in E.coli by using gene recombination technique. The fused protein is purified preliminarily.