狂犬病病毒糖蛋白基因中和抗原表位在大肠杆菌中的串联表达

Expression of neutralizing antigenic determinants of rabies virus glycoprotein in tandem in Escherichia coli

采用RT-PCR方法从狂犬病病毒HEP株中克隆狂犬病糖蛋白膜外区全长基因。利用突变PCR方法,将糖蛋白基因中三段中和抗原位点串联,克隆至pET-28a表达载体。阳性重组质粒转化大肠杆菌BL21(DE3)后,用IPTG诱导表达并确定最佳诱导条件。SDS-PAGE电泳结果显示所表达的蛋白大小与预期相符。经Western-blotting检测,串联表达的狂犬病糖蛋白基因中和抗原位点可与狂犬病标准阳性血清发生特异性反应,表明重组蛋白具有良好的反应原性。

The gene fragment encoding the extracellular domain of rabies virus （RV） glycoprotein was amplified from RV （HEP strain）. Three neutralizing antigenic determinants of glycoprotein were spliced together in tandem using mutation PCR and were inserted into PET-28a vector. The recombinant plasmids were transformed into E. coli BL21 （DE3） competent cells for expression. The recombinant protein was optimized with proper induction by IPTG. SDS-PAGE electrophoresis analysis revealed a band of protein correspondent with molecular weight of the target protein. The recombinant protein was recognized specifically by anti-RV positive serum in western-blotting analysis.