猪瘟病毒T细胞表位与猪细小病毒VP2融合基因在重组杆状病毒中的表达

Expression of fusion gene consisting of T cell antigen epitope gene of classical swine fever virus and VP2 protein gene of porcine parvovirus in recombinant baculovirus

将猪细小病毒分离株LJL12 VP2基因和猪瘟病毒多肽基因E290克隆至真核表达载体pMel Ba-cA,将该重组质粒与Bac-N-Blue DNA共转染昆虫细胞,并对表达产物进行分析,构建了表达猪瘟病毒T细胞表位和猪细小病毒VP2融合蛋白的重组杆状病毒。结果显示,获得了含VP2基因和E290基因的重组质粒pM-VP2-E290,昆虫细胞sf9的表达产物经SDS-PAGE、Western-blot检测后确定所表达的蛋白质分子质量大小约为67 ku,且具有天然蛋白的抗原特性。免疫电镜观察可见表达产物形成的病毒样颗粒。证实,成功构建了同时含有PPV VP2基因和CSFV特异性T细胞表位基因的重组杆状病毒,并在昆虫细胞中得到了高效表达。

To construct recombinant baculovirus expressing fusion gene of T cell antigen epitope gene of classical swine fever virus（CSFV） with VP2 protein gene of porcine parvovirus （PPV） ,the VP2 gene of PPV isolate LJL12 and E290 gene of CSFV were co-cloned into pMel BacA vector. Then,the recombinant plasmid and Bac-N-Blue DNA were co-transfected into sf9 cells. The expressed fusion protein was analyzed by SDS-PAGE and Western-blotting. The expressed product which was approximately 67 ku in size reacted strongly with the PPV-specific antibody. The virus-like particles formed by expressed product were observed using an immunoelectron microscopy. The results confirmed that the recombinant baculovirus containing T cell antigen epitope gene of CSFV and VP2 protein gene of PPV was constructed successfully and expressed efficiently in sf9 cells.