猪流感病毒抗原表位基因表达载体构建与免疫原性分析.pdfVIP

生 物 工 程 学 报 Chin J Biotech 2008, April 25; 24(4): 690-694 Chinese Journal of Biotechnology ISSN 1000-3061 cjb@ © 2008 Institute of Microbiology, CAS CSM, All rights reserved 研究简报 刘惠莉, 邢继兰, 潘洁, 杨秋峰, 赵艳敏 , 201106 : 根据猪流感病毒血凝素蛋白基因(Heamuglutinine, HA)的核苷酸序列, 设计、筛选HA 蛋白氨基酸序列的主要表 位多肽4 个, 将4 个片段以柔性连接串联成模拟蛋白, 核苷酸约为 300 bp, 体外扩增该模拟蛋白基因, 插入到原核表达 载体pET30a(+) 中, 转染宿主菌诱导表达, 结果获得分子量为20 kD 的表达蛋白, 该蛋白可与抗His-tag 抗体、抗猪流感 病毒H1N1 、H3N2 亚型高免血清发生免疫学反应。纯化后免疫小鼠, ELISA 及血凝抑制(Heamuglutinine inhibitor, HI)试 验检测, 小鼠产生针对多肽抗原的血清抗体, 同时还可检测到H1N1 、H3N2 亚型 SIV 血凝抗体。流氏细胞仪检测免疫 组外周血淋巴细胞高于对照组, 说明该模拟蛋白具有与 H1N1 、H3N2 亚型猪流感病毒相似的免疫原性及反应原性, 为 H1N1 、H3N2 血清亚型猪流感病毒疫苗研制提供了新手段。 : 猪流感病毒, 抗原表位, 新型疫苗 Construction and Immunogenicity Analysis of Antigenic Epitopes of Swine Influenza Virus Huili Liu, Jilan Xing, Jie Pan, Qiufeng Yang, and Yanmin Zhao Institute of Animal Science and Veterinary Medicine , Shanghai Academy of Agriclutural Scinece, Shanghai 201106, China Abstract: Several antigen epitopes were designed according to the sequences of Swine influenza virus hemagglutinin (HA) genes and lined with the interval. The gene was amplified by PCR and sub cloned into pET30a (+) vector. The fusion protein was expressed in E. coli BL21 (DE3) by induced with IPTG and purified by affinity chromatography. The molecular weight of the protein was about 20 kD in SDS. Immunological activity of the fusion protein was analyzed by Western blot. The results showed that the fusion protein could interact with anti-His antibody and the rabbit antiserum against SIV. The immunized mouse can produced antibodies against the target peptide and HI antibody against SIV H1N1 or H3N2. This stud