猪流行性腹泻病毒主要结构蛋白在杆状病毒中的表达及免疫原性分析

Expression of major structural protein of pig epidemic diarrhea virus in baculovirus and analysis of its immunogenicity

为了研制一种有效控制猪流行性腹泻的疫苗,本试验设计了两对引物,分别扩增猪流行性腹泻病毒(PEDV)主要抗原表位S1(S蛋白中一个有效的抗原位点)和M蛋白的基因。将两段基因克隆到杆状病毒双表达载体pFastBacTMDuai中,构建了重组转移质粒pFBD-S1-M,并将其转化至DH10BacTM感受态细胞中制备重组穿梭质粒Bacmid-S1-M,转染Sf9昆虫细胞后,拯救出能够共表达S1和M蛋白的重组杆状病毒rBacmid-S1-M。表达产物经过SDS-PAGE和Western blot检测,共表达的重组S1和M蛋白产物能够与PEDV多克隆抗体发生特异性反应,表明共表达的蛋白具有良好的反应原性。本研究为PEDV基因工程亚单位疫苗的研制奠定了基础。

To develop an effective vaccine against pig epidemic diarrhea,the genes of major antigen epitope S1（an antigenic determinant of S protein） and M protein of porcine epidemic diarrhea virus（ PEDV） were amplified by PCR,respectively. The PCR products were cloned into baculovirus double expression vector pFastBacTMDuai and transformed into DH10Bac- host cells to produce the recombinant shuttle plasmid Bacmid- S1- M. Then the shuttle plasmid was used to transfect Sf9 insect cells to get recombinant baculovirus rBacmid- S1-M. The protein expression was demonstrated by SDS- PAGE and western blot. The results showed that the recombinant product of PEDV S1 and M proteins can specifically react with PEDV polyclonal antibody,indicating that the expressed protein has good antigenicity. This study provides a good basis for the development of subunit vaccine against PEDV.