猪瘟病毒E2基因噬菌体展示多肽库的构建及表位鉴定

Construction of Phage Display Libraries of Classic Swine Fever Virus E2 Gene and Identification of E2 Protein Epitopes

本研究通过建立CSFV噬菌体展示多肽库,并对其进行生物淘选,以期获得E2蛋白上新的抗原表位。选择CSFV石门株(SM)和疫苗株(HCLV)为代表株,采用噬菌体展示技术,以T7select415-1b为载体,分别构建了CSFV SM株和HCLV株E2基因噬菌体展示多肽库(SM-E2库和HCLV-E2库)。通过生物淘选和噬菌体原位杂交技术,采用7株猪瘟单克隆抗体和1株猪瘟高免血清分别对构建的SM-E2库和HCLV-E2库进行抗原表位筛选。结果共筛选到了5条与E2蛋白高度同源的序列,在E2蛋白上的同源区域分别为TAVSPTTLR、YYEP、TTWKEYSH、GGQ(V)VK和PDGLPHY。结果表明,TAVSPTTLR、YYEP和TTWKEYSH序列与目前已知E2蛋白表位一致,说明它们是E2蛋白上的优势表位;GGQ(V)VK和PDGLPHY序列与预测表位一致,推测是E2蛋白上潜在的抗原表位。

The phage display peptides libraries of Classic Swine fever virus（CSFV）E2 gene were constructed to identify the potential epitopes of E2 protein by biopanning.The E2 genes of Shimen strain（SM） and hog cholera lapinized virus（HCLV） strain were cloned into T7seclect-415b vector respectively.So the E2 gene phage display peptides libraries of Shimen strain and HCLV strain were constructed and named as SM-E2 libraries and HCLV-E2 libraries.CSFV polyclonal antibody（PcAB） and seven strains of McAbs were used to screen the epitopes in SM-E2 libraries and HCLV-E2 libraries by biopanning and phage in situ hybridization.After analyzing the sequences of selected phage recombinants,five sequences which were highly homologous with TAVSPTTLR,YYEP,TTWKEYSH,GGQ（V）VK and PDGLPHY were selected by the MAbs and CSFV PcAB.These results proved that these three sequences of TAVSPTTLR,YYEP and TTWKEYSH are dominant epitopes on E2 protein,which were reported in the previous papers.In addition,two sequences of GGQ（V）VK and PDGLPHY corresponds to the epitopes predicted by computer.So these two sequences might be the potential epitopes of E2 protein.