中国农业科学 ›› 2008, Vol. 41 ›› Issue (5): 1503-1510 .doi: 10.3864/j.issn.0578-1752.2008.05.033

• 兽医·资源昆虫 • 上一篇    下一篇

含CpG序列PRRSV ORF5基因真核质粒构建及其免疫原性的研究

温建新,李 俊,周 顺,任慧英,刘文华,邹 玲,徐 文,牛钟相,王金宝,吴家强   

  1. 山东农业大学动物科技学院
  • 收稿日期:2007-11-22 修回日期:2008-02-01 出版日期:2008-05-10 发布日期:2008-05-10
  • 通讯作者: 王金宝

Studies of immunogenicity responses of mice immuned with eukaryon expression plasmid on PRRSV SD2 E containing CpG motifs

  

  1. 山东农业大学动物科技学院
  • Received:2007-11-22 Revised:2008-02-01 Online:2008-05-10 Published:2008-05-10

摘要: 【目的】提高猪繁殖与呼吸综合征病毒(PRRSV)基因免疫的效果,构建含CpG基序和PRRSV ORF5的真核重组表达质粒CpG-pVAX1-ORF5。【方法】经酶切鉴定和序列测定,将重组质粒转染COS-7细胞,经间接免疫荧光试验证实CpG-pVAX1-ORF5可表达PRRSV GP5蛋白。免疫SPF小鼠,检测鼠的特异性抗体滴度、脾T淋巴细胞亚群数量(CD4+和CD8+)以及淋巴细胞增殖反应(MTT法)水平。【结果】本试验构建的含CpG基序真核重组表达质粒CpG-pVAX1-ORF5能诱导小鼠产生针对PRRSV的体液免疫与细胞免疫。【结论】CpG-ODN可提高PRRSV基因疫苗的免疫效果。

关键词: 猪繁殖与呼吸综合征病毒, ORF5基因, DNA免疫

Abstract: The recombinant expression vector plasmid CpG-pVAX1-E was transfected into COS-7 cells by using lipofectamine methods.The ORF5 mRNA of transfected cells were detected by RT-PCR. The transient expression of PRRSV SD2 ORF5 nucleocapsid protein was detected by indirect immunofluorescence assay(IFA). In some transfected COS-7 cells, the green fluorescence was showed.The recombinant expression plasmid was inoculated into SPF mice.The antibody ies specific for E protein in serum were confirmed by ELISA. T lymphocyte proliferation test(MTT) were used to detect the number of CD4+ 、CD8 + and the T lymphocyte proliferation in peripheral blood of mice vaccinated. The results showed that the recombinant plasmid induced CpG-pVAX1-E could induce higher response of cellular immune response and specific humoral immune responses in mice . CpG-ODN was effective adjuvant which could significantly enhance the immunogenicity of PRRSV SD2 ORF5 DNA vaccine.

Key words: Porcine reproductive and respiratory syndrome virus(PRRSV)SD2, Eukaryotic expression vector, E Gene, Immunogenicity