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猪流行性腹泻病毒(porcine epidemic diarrhea virus,PEDV)纤突蛋白S1能够介导宿主抵御PEDV感染的中和抗体的产生,是PEDV基因工程疫苗研究的重要靶抗原。将PEDV的S1基因克隆至杆状病毒基因组中,构建包含PEDV S1基因的重组杆状病毒,使S1蛋白表面展示表达。以该重组病毒作为活载体疫苗皮下接种BALB/c小鼠,验证S1蛋白在重组杆状病毒中的表达及其免疫原性。Western blotting分析被重组杆状病毒感染的Sf9细胞中,PEDV S1蛋白能有效表达且呈现大小分别约120 k D和150 k D的2条带,其中前者至少比预期大30 k D左右,推测重组S1蛋白在Sf9细胞中被高度糖基化修饰。重组杆状病毒的胶体金免疫电子显微镜观测结果显示S1蛋白展示于杆状病毒表面;间接ELISA试验结果证明重组杆状病毒可以诱导产生PEDV特异性抗体,抗体效价达到1∶5 000。血清中和试验及淋巴细胞增殖试验结果证明,该重组杆状病毒能够激发有效的体液免疫及细胞免疫反应,S1蛋白在重组杆状病毒表面展示表达并具有免疫原性。
Porcine epidemic diarrhea virus (PEDV) splicing protein S1 can mediate the host immune response against PEDV infection, which is an important target antigen for PEDV genetic engineering vaccine research. The S1 gene of PEDV was cloned into the baculovirus genome to construct a recombinant baculovirus containing the PEDV S1 gene to express the surface of the S1 protein. BALB / c mice were inoculated subcutaneously with this recombinant virus as a live vector vaccine to verify the expression of S1 protein in recombinant baculovirus and its immunogenicity. Western blotting analysis of recombinant baculovirus Sf9 infected cells, PEDV S1 protein was effectively expressed and showed the size of about 120 kD and 150 kD 2 bands, of which the former at least 30 kD larger than expected, presumably recombinant S1 protein is highly glycosylated in Sf9 cells. The recombinant baculovirus showed that the S1 protein was displayed on the surface of baculovirus. The results of indirect ELISA showed that the recombinant baculovirus could induce the production of PEDV-specific antibody. The antibody titer reached 1: 5000. Serum neutralization test and lymphocyte proliferation test results show that the recombinant baculovirus can stimulate effective humoral and cellular immune response, S1 protein expressed on the surface of recombinant baculovirus and is immunogenic.