中文摘要：

目的克隆人附睾蛋白酶抑制蛋白(epididymal protease inhibitor,Eppin)基因,构建原核表达载体,诱导其在大肠杆菌中表达,并鉴定重组蛋白的免疫学活性。方法采用RT-PCR克隆人Eppin基因,将其克隆到原核表达载体pMAL-c2X,并通过IPTG诱导目的基因在大肠杆菌中表达。Amylose树脂预装柱层析法提纯重组蛋白,并用SDS-PAGE和蛋白印迹法鉴定。结果PCR扩增出396bp目的基因片段Eppin。工程菌pMAL-c2X-Eppin经IPTG诱导表达后,SDS-PAGE显示有新生的蛋白表达条带,Mr约为54000与预期的一致,重组蛋白用Amylose树脂提纯,纯化蛋白的蛋白质经SDS-PAGE分析可见单一条带。Western blot显示重组蛋白质有良好的免疫活性。结论获得了Eppin融合蛋白在原核系统中的稳定表达,为后续深入研究其抗生育功能奠定基础。

英文摘要：

Objective To construct prokaryotic expression vector for human Eppin gene and analyze the characteristics of expressed fusion protein. Methods Human Eppin cDNA was obtained from human testis tissue through RT-PCR,and then cloned into the pMAL-c2X vector. The recombinant protein was achieved in E.coli through IPTG induction. The expressed product was purified with Amylose resin,while the immunological activity of the expression product was detected by Western blotting. Results The gene fragment with length o...