牛皮蝇HypoderminA基因在大肠杆菌中的表达

Expression of hypodermin A gene of H. lineatum in Escherichia coli

从牛皮蝇Ⅱ期幼虫中提取总RNA,进行RT-PCR扩增牛皮蝇Hypodermin A基因。将扩增基因进行克隆测序,构建原核表达载体pET30-HA,转化大肠杆菌BL21(DE3)。用IPTG诱导表达,得到表达量达到全菌蛋白35%以上的特异性蛋白。SDS-PAGE分析表明,表达产物大部分以包涵体形式存在。Western-blot检测结果表明,获得的重组蛋白为重组Hypodermin A,具有较好的免疫活性。

Total RNA was extracted from the second stage larva of H. lineatum. Hypodermin A gene of H. lineatum was amplified by RT-PCR. The target gene was cloned and sequenced. The prokaryotic expression vector pET30-HA was constructed and then transformed to E. coli BL21 （DE3）. The recombinant E. coli was induced and expressed by IPTG. The expression level of the specific protein was more than 35% of total protein. It was mainly present in inclusion body observed by SDS-PAGE. Western blot analysis indicated that the obtained protein was recombinant hypodermin A,and had good reactiogenicity against positive serum.