禽多杀性巴氏杆菌C_(48-3)株成熟黏附蛋白的表达及其免疫原性检测  

Expression of Mature Adhesive Protein of Avian Pasteurella multocida C_(48-3)and Detection of its Immunogenicity

作　　者：严芳[1] 何翠[1] 张磊[1] 吾鲁木汗·那孜尔别克[1] 罗南书[1] 恩特马克·布拉提白[1] 

机构地区：[1]吉首大学生物资源与环境科学学院,部省共建生物工程实验室,湖南吉首416000

出　　处：《生物技术通讯》2009年第6期823-826,共4页Letters in Biotechnology

基　　金：吉首大学研究生教育教学改革研究项目(2006011)

摘　　要：目的:在大肠杆菌原核表达系统中高效表达禽多杀性巴氏杆菌成熟黏附蛋白Cpm39,并检测其免疫原性。方法:通过BamHⅠ和SalⅠ双酶切重组载体pMD18-cpm39获得cpm39基因片段,将该基因片段克隆至表达载体pMAL-p2X上,构建表达质粒pMAL-p2X-cpm39,转化至大肠杆菌BL21(DE3),在IPTG诱导下表达融合蛋白,用禽多杀性巴氏杆菌C48-3株Cp39天然粘附蛋白的免疫血清经Western印迹检测其免疫原性。结果:SDS-PAGE结果显示表达的融合蛋白相对分子质量为78×103,与预期结果相符,而Western印迹结果表明诱导表达的融合蛋白MBP-Cpm39能与Cp39天然黏附蛋白抗体发生特异性反应。结论:构建了表达质粒pMAL-p2X-cpm39,获得了具有免疫原性的重组融合蛋白,为进一步研究禽多杀性巴氏杆菌成熟黏附蛋白的免疫保护功能奠定了基础。Objective： To express the recombinant mature adhesive protein Cpm39 of avian Pasteurella multocida strain C4s-3 and detect its immunogenicity. Methods： The cpm39 gene was cloned into the expression vector pMAL-p2X and constructed recombinant plasmid pMAL-p2X-cpm39. The pMAL-p2X-cpm39 was introduced into E.coli BL21 （DE3） by transformation, and the recombinant MBP-Cpm39 was expressed in E.coli after IPTG inducing. The fusion protein of MBP- Cpm39 was detected by using SDS-PAGE and Western blots. Results： SDS-PAGE showed that the molecular weight of 78x103 recombinant protein was expressed in E.coli with IPTG inducing. In Western blot analysis, mice antiserum against purified native Cp39 reacted with the MBP-Cpm39, it demonstrated that the recombinant protein is a immunogenic protein. Conclusion： Recombinant plasmid pMAL-p2X-cpm39 was successfully constructed and the target protein was expressed in E.coli. The recombinant protein might be a useful vaccine candidate antigen for avian P.multocida.

关 键 词：禽多杀性巴氏杆菌 成熟黏附蛋白 原核表达 免疫原性 

分 类 号：Q78[生物学—分子生物学] R392.1[医药卫生—免疫学]