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新城疫强毒株F蛋白裂解位点串联基因的构建及其原核表达
引用本文:刘成宏,金扩世,金宁一,鲁会军,贾雷立,陈义锋,金明兰,姜鲲.新城疫强毒株F蛋白裂解位点串联基因的构建及其原核表达[J].粉末涂料与涂装,2007,20(2):73-76.
作者姓名:刘成宏  金扩世  金宁一  鲁会军  贾雷立  陈义锋  金明兰  姜鲲
作者单位:吉林大学农学部畜牧兽医学院 长春130062(刘成宏,陈义锋,金明兰,姜鲲),军事医学科学院军事兽医研究所 长春130062(金扩世,金宁一,鲁会军,贾雷立)
基金项目:吉林省科技计划发展项目
摘    要:目的构建鸡新城疫(ND)强毒株多裂解位点串联基因表达载体,为建立NDV强弱毒株的ELISA鉴别方法奠定基础。方法人工合成一段包含4种不同NDV强毒株裂解位点的基因Fe(F prote in multitude epi-position),其中各裂解位点之间用Linker(GGGGS)使其串联,然后进行2倍和4倍拷贝,并定向克隆至原核表达质粒pET-28a(+)中,构建重组表达质粒pET-28a(+)/2Fe和pET-28a(+)/4Fe,进行表达和Western blot鉴定。结果构建的原核表达重组质粒pET-28a(+)/2Fe和pET-28 a(+)/4Fe测序正确。Western blot检测结果表明,2Fe、4Fe蛋白均与新城疫强毒株F48E9阳性血清呈阳性反应,而与新城疫弱毒株V4阳性血清呈阴性反应。结论已成功构建NDV强毒株F蛋白裂解位点串联基因表达载体,为建立NDV强弱毒株的ELISA鉴别方法提供理论依据。

关 键 词:新城疫  强毒株  裂解位点  串联基因
文章编号:1004-5503(2007)02-073-04
收稿时间:2006-09-01
修稿时间:2006年9月1日

Construction of Prokaryotic Expression Vector for F Protein Schizolysis Site Gene Cascade of Virulent Newcastle Disease Virus Strain
LIU Cheng-hong ,JIN Kuo-shi,JIN Ning-yi,et al.Construction of Prokaryotic Expression Vector for F Protein Schizolysis Site Gene Cascade of Virulent Newcastle Disease Virus Strain[J].Chinese Journal of Biologicals,2007,20(2):73-76.
Authors:LIU Cheng-hong  JIN Kuo-shi  JIN Ning-yi  
Affiliation:College of Animal Husbandry and Veterisity,Jilin University, Changchun 130062, China
Abstract:Objective To construct the prokaryotic expression vector for F protein schizolysis site gene cascade of virulent Newcastle disease virus(NDV) strain and lay a foundation of differentiation of virulent and low virulent NDV strains by ELISA.Methods Synthesize a gene fragment Fe(F protein multitude epi-position) containing 4 different schizolysis sites of virulent NDV strains,of which the sites were linked through a linker GGGGS.Copy Fe gene for 2 and 4 times,then clone the obtained 2Fe and 4Fe genes directly into prokaryotic expression vector pET-28a( ) respectively.Transform the constructed recombinant plasmids pET-28a( )/2Fe and pET-28a( )/4Fe to E. coli BL21(DE3) respectively and identify the expressed products by Western blot.Results Sequencing results proved that recombinant plasmids pET-28a( )/2Fe and pET-28a( )/4Fe were correctly constructed.Western blot showed specific reaction of expressed 2Fe and 4Fe proteins with F48E9 positive sera but no reaction with V4 positive sera.Conclusion A prokaryotic expression vector for F protein schizolysis site gene cascade of virulent NDV strain was successfully constructed,which provided a theoretical basis for differentiation of virulent and low virulent NDV strains by ELISA.
Keywords:Newcastle disease  Virulent strain  Schizolysis site  Gene cascade
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