猪脑心肌炎病毒GXLC株VP1基因的原核表达及鉴定

Prokaryotic Expression and Identification of VP1 Gene of Encephalomyocarditis Virus GXLC Strain from Swine

根据猪脑心肌炎病毒(EMCV)GXLC株的基因组序列设计一对特异性引物,应用RT-PCR方法扩增EMCV VP1基因目的片段,将其克隆至原核表达载体pET-32a(+),构建了EMCV VP1基因重组表达质粒pET-32a-VP1。将pET-32a-VP1转化BL21(DE3)株感受态细胞,并用IPTG进行诱导表达。结果显示,经SDS-PAGE分析,VP1蛋白获得高效表达,融合蛋白质的分子质量约为52 ku,表达的重组蛋白以包涵体形式存在。经Western blot,结果显示所表达的VP1蛋白与猪抗EMCV阳性血清发生特异性的免疫印迹反应,表明其具有良好的抗原反应原性。

One pair of primers were designed according to the genomic sequence of porcine encephalomyocarditis virus（EMCV）GXLC strain,and the VP1 gene was amplified by RT-PCR.The target fragment was cloned into prokaryotic expression vector pET-32a（＋） to construct a recombinant expression vector pET-32a-VP1.Then,the pET-32a-VP1 plasmid was transformed into E.coli BL21（DE3）strain and induced by IPTG to produce recombinant VP1 protein.The results of SDS-PAGE analysis showed that the recombinant VP1 protein was efficiently expressed in the form of inclusion body and the molecular weight of the fusion protein was about 52 ku.The results of Western blot test revealed that the purified VP1 protein had a specific reaction with pig anti-EMCV positive serum,indicating that it had good antigenicity.