羊传染性脓疱病毒新疆流行株F1L基因克隆及表达  

Cloning and Expression of F1L Gene of Orf Virus in Xinjiang

作　　者：李瑞芳[1] 乔军[1] 张辉[1] 陈创夫[1] 

机构地区：[1]石河子大学动物科技学院,新疆石河子832000

出　　处：《生物技术》2012年第3期13-16,共4页Biotechnology

基　　金：国家"973"计划项目(2010CB30203);国际科技合作项目(2006DFA33740)资助

摘　　要：目的:为了探讨F1L重组蛋白作为羊传染性脓疱病毒亚单位疫苗的可行性。方法:以分离出羊传染性脓疱病毒ORFV-shz分离株为模板,扩增F1L基因片段,并将其克隆至pMD18-T载体中进行测序。构建原核表达载体pET-32a-F1L,转化至大肠埃希菌BL21,用SDS-PAGE及Western blot检测目的蛋白的表达情况及其反应原性。结果:PCR扩增出F1L基因全长1023bp,共编码340个氨基酸;推导的氨基酸序列中第1～70位氨基酸构成信号肽序列,第285～313位氨基酸为跨膜区。SDS-PAGE分析表明获得了约57.4kDa的融合蛋白,在Western-blotting检测中,融合蛋白可与羊传染性脓疱病毒阳性血清发生特异性反应,表明其具有良好的反应原性。结论:成功构建F1L基因的原核表达载体,为研究新疆地区羊传染性脓疱的防治及疫苗开发提供了科学依据。Objective:In order to investigate the viability of F1L recombinant proteins as contagious ecthyma virus subunit vaccine.Method:The F1L gene fragment was amplified from the genome of ORFV-shz strain and ligatured with pMD18-T vector for sequencing.The recombinant prokaryotic expression vector pET-32a-F1L was constructed,and transformed into Escherichia coli BL21.The expressed protein was analyzed by SDS-PAGE,and the reactinoge-nicity of the protein was detected by Western blot.Result:The F1L gene was 1023bp,which encoded 340 amino acids.The front 70 amino acids constitute signal peptide,and 285-313 amino acids from transmembrane.Target protein analyzed by SDS-PAGE was about 57.4kDa in side.Western-blotting showed the protein could specifically interact with ORFV positive serum,indicating that the protein had good reactinogenicity.Conclusion:The recombinant prokaryotic expression vector pET-32a-F1L was successfully constructed,and our study should be viewed as a new window of opportunity in prevention and vaccine development of Orf in Xinjiang area.

关 键 词：羊传染性脓疱病毒 F1L基因 序列分析 原核表达 反应原性 

分 类 号：S852.65[农业科学—基础兽医学]