猪圆环病毒2型ORF2基因重组植物乳杆菌的构建及其免疫原性分析

发布时间：2018-05-16 22:14

本文选题：猪圆环病毒型 + Cap蛋白　；参考：《中国畜牧兽医》2017年08期

【摘要】：为研制一种能有效控制猪圆环病毒2型(porcine circovirus type 2,PCV2)的新型疫苗,本试验将PCV2的ORF2基因克隆至植物乳杆菌表达载体pSCPSP超强启动子SCP的下游,构建重组表达质粒pSCPSP-Cap,用电击转化法转化克隆至表达宿主菌植物乳杆菌,获得一株重组植物乳杆菌。以SDS-PAGE和Western blotting法检测24h上清液中表达的蛋白;将8周龄的SPF级BALB/c小鼠随机分组,每组8只,分别设置对照组、空载体组、灌胃组、饮水组、灭活疫苗组,试验14、28、42和56d后对小鼠采血,分离血清,利用PCV2ELISA抗体检测试剂盒检测小鼠血清中PCV2的抗体水平。SDS-PAGE、Western blotting结果显示,PCV2衣壳蛋白在植物乳杆菌中成功分泌表达,表达的蛋白分子质量为28ku,且具有良好的反应原性,重组植物乳杆菌免疫小鼠诱导机体产生PCV2特异性抗体显著高于其他组(P0.05)。结果表明,PCV2的ORF2基因在植物乳杆菌中成功表达,且具有良好的生物活性,可为PCV2口服疫苗的开发研究提供理论参考。
[Abstract]:In order to develop a novel vaccine for the control of porcine circovirus type 2 PCV2 (porcine circovirus type 2), the ORF2 gene of PCV2 was cloned into the downstream of pSCPSP superpromoter SCP, an expression vector of Lactobacillus plantarum. The recombinant expression plasmid pSCPSP-Cap. was constructed and cloned into the host bacterium Lactobacillus plantarum by electroporation method, and a recombinant Lactobacillus plantarum strain was obtained. The expression of protein in the supernatant of 24 hours was detected by SDS-PAGE and Western blotting, and 8 weeks old SPF grade BALB/c mice were randomly divided into control group, empty carrier group, gastric perfusion group, drinking water group and inactivated vaccine group. Blood samples were collected from mice after 14 days and 56 days later. The antibody level of PCV2 in mouse serum was detected by PCV2ELISA antibody detection kit. The results of SDS-PAGEG Western blotting showed that capsid protein of PCV2 was secreted and expressed successfully in Lactobacillus plantarum. The molecular weight of the expressed protein was 28ku. and the protein had good reactivity. The PCV2 specific antibody produced by recombinant Lactobacillus plantarum immunized mice was significantly higher than that of other groups. The results showed that the ORF2 gene of PCV2 was successfully expressed in Lactobacillus plantarum and had good biological activity, which could provide a theoretical reference for the development and study of oral PCV2 vaccine.
【作者单位】：华南农业大学兽医学院广东省动物源性人畜共患病预防与控制重点实验室国家生猪种业科学技术研究中心;宜春学院;
【基金】：广东省省级科技计划项目(2013B020202002)
【分类号】：S852.651

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