小反刍兽疫病毒H和N蛋白的表达及抗体制备  

Expression and antibody preparation of H and N proteins of the Peste des petits ruminants virus

作　　者：孟庆玲[1,2] 乔军[1] 倪伟[1] 才学鹏[2] 骆学农[2] 张娜[1] 

机构地区：[1]石河子大学动物科技学院预防兽医学重点实验室,新疆石河子832003 [2]中国农业科学院兰州兽医研究所家畜疫病病原生物学国家重点实验室,甘肃兰州730046

出　　处：《中国病原生物学杂志》2010年第9期645-648,644,共5页Journal of Pathogen Biology

基　　金：家畜疫病病原生物学国家重点实验室开放课题(No.KEYLA200802);石河子大学高层次人才专项(No.RCZX200801)

摘　　要：目的表达小反刍兽疫病毒（PPRV）疫苗株血凝蛋白（H）和核蛋白（N）并制备其抗体。方法利用RT-PCR技术扩增PPRV H和N蛋白基因进行序列分析,并分别克隆入pET-28a（＋）原核表达载体中,构建重组载体pETH和pETN;重组载体分别转化E.coli BL21（DE3）,用IPTG诱导表达,SDS-PAGE和Western blot分析表达产物。结果 PPRV疫苗株H和N基因ORF全长分别为1 830 bp和1 578 bp,分别编码609个和525个氨基酸。经IPTG诱导6 h后,H和N蛋白基因片段在大肠埃希菌中均获得了高效表达,表达的重组蛋白均具有良好的抗原性。分别用纯化的H和N重组蛋白免疫昆明系小鼠,获得的抗体效价分别为1∶16~1∶32和1∶32~1∶64。结论成功表达了PPRVH和N蛋白并制备了相应抗体,为研发PPRV检测试剂奠定了基础。Objective To clone hemagglutinin（H） protein and nucleoprotein（N） genes of a Peste des petits ruminants virus（PPRV） vaccine strain and prepare their antibodies.Methods H and N genes of PPRV were amplified and sequenced and then cloned into pET-28a（＋） to generate recombinant vectors pETH and pETN for expression.Transformed E.coli BL21（DE3）was analyzed by SDS-PAGE and Western blot.Results The open reading frame length of H and N genes of the PPRV vaccine strain were 1 830 bp and 1 578 bp and encoded 609 and 525 amino acids,respectively.Fragments of H and N genes were expressed in recombinant E.coli BL21（DE3）6 h after induction of IPTG,which had better reactogenicity.High titers of antibodies against recombinant H and N proteins were successfully prepared by immunization of Kunming mice.Conclusion H and N proteins and their antibodies were successfully produced,laying the foundation for agents to detect PPRV infection.

关 键 词：小反刍兽疫病毒 血凝蛋白 核蛋白 表达 抗体制备 

分 类 号：S852.65[农业科学—基础兽医学]