番鸭呼肠孤病毒ZJ99株σC基因的序列分析及其在大肠杆菌中的表达

参考已发表的番鸭呼肠孤病毒(Muscovy duck reovirus, mDRV)的S4序列，自行设计1对扩增σC基因的引物，对mDRV ZJ99株的RNA抽提物进行RT-PCR扩增，成功地获得了843 bp特异性的扩增片段(GenBank: AY619690)。将PCR产物克隆测序，BLASTn比较结果显示, mDRV ZJ99株σC基因与福建MW9710株对应基因的同源性为98％，与法国89026和89330株对应基因的同源性分别为92％和93%； BLASTp比较结果显示, ZJ99株编码的σC蛋白与MW9710、89026和89330株相应蛋白分别具有94％、89％、 89％同一性(identity)和95％、92％、93％相似性(similarity)。进一步将σC基因亚克隆至原核表达载体pET28a， 转化大肠杆菌(Escherichia coli )BL21(DE3)，IPTG诱导阳性重组子，SDS-PAGE电泳检测发现, σC蛋白在大肠杆菌中以包涵体的形式高效表达，目的蛋白的表达量可达到细菌总蛋白的22.9％。

Sequence Analysis of the σC Gene of Muscovy duck reovirus ZJ99 Strain and Its Expression in Escherichia coli

The σC gene (GenBank: AY619690) of Muscovy duck reovirus (mDRV) ZJ99 strain was amplified and sequenced with RT-PCR. BLASTn analysis results showed that the σC gene of mDRV ZJ99 had 98% homology to mDRV MW9710 isolated in Fujian Province of China, 92% and 93% homology to mDRV 89026 and mDRV 89330 reported in France. BLASTp analysis showed that the deduced amino acid sequence of ZJ99 had 94%, 89%, 89% identities and 95％, 92％, 93％ similarities to that of MW9710, 89026 and 89330, respectively. The σC gene was further subcloned into pET-28a and transformed into Escherichia coli BL21(DE3). SDS-PAGE analysis showed that the σC protein was successfully expressed in E. coli, which existed as inclusion body and accumulated up to 22.9% of total cellular proteins.