含RGD受体结合位点口蹄疫病毒Asia1/JS/China/2005株的拯救及初步鉴定  

Rescue and identification foot-and-mouth disease virus Asia1/JS/China/2005 strain with Arg-Gly-Asp RGD receptor recognition site

作　　者：李平花[1] 白兴文[1] 曹伟军[2] 卢曾军[1] 孙普[1] 殷宏[1] 刘在新[1] 

机构地区：[1]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,农业部畜禽病毒学重点开放实验室,国家口蹄疫参考实验室,兰州730046 [2]甘肃农业大学动物医学院,兰州730070

出　　处：《微生物学报》2009年第7期943-948,共6页Acta Microbiologica Sinica

基　　金：国家支撑计划(2006BAD06A12);国家”973”项目(2005CB523201)~~

摘　　要：【目的】构建含有精氨酸-甘氨酸-天冬氨酸(RGD)受体结合位点口蹄疫病毒(FMDV)Asia1/JS/China/2005株的全长感染性cDNA克隆。【方法】采用定点突变方法,构建Asia1型FMDV含有预期突变的全长cDNA克隆pFMDV-RGD。pFMDV-RGD重组质粒经NotI线化后,与表达T7 RNA聚合酶的真核质粒pcDNAT7P共转染BHK-21细胞,进行FMDV-RGD病毒拯救。【结果】序列测定结果表明成功构建了FMDV含有RGD受体位点的Asia1/JS/China/2005全长cDNA克隆。共转染试验获得拯救病毒,对拯救的病毒分别进行序列测定、间接免疫荧光、电子显微镜观察和乳鼠致病性分析,表明成功拯救了含有RGD受体结合位点的Asia1/JS/China/2005株FMDV。【结论】该试验为进一步研究含有RGD和RDD受体结合位点2个拯救病毒生物学特性的差异奠定了基础。[Objective] To construct an infectious full-length cDNA clone of Asial/JS/China/2005 strain with Arg-Gly-Asp （RGD） receptor recognition site. [ Methods]We constructed foot-and-mouth disease virus type Asial full-length cDNA clone pFMDV-RGD by using site-directed mutagenesis. The plasmid pFMDV-RGD contained the desired mutation. The plasmids pFMDV-RGD were linearized with NotI enzyme. Linearized plasmid and pcDNATTP plasmids expressing T7 RNA polymerase cotransfected into BHK-21 cells to rescue FMDV-RGD. [ Results] We constructed FMDV Asial/JS/China/2005 strain full-length cDNA clone with Arg-Gly-Asp receptor recognition site by sequence. We obtained rescued virus by plasmid cotransfection. The results of sequencing, indirect immunofluorescence, electron microscope and sulk mice pathogenicity analysis showed foot-and- mouth disease virus containing Arg-Gly-Asp receptor recognition site was successfully rescued. [ Conclusion] The results lay a foundation for further study of biology characteristic diversity of rescued virus with Arg-Gly-Asp and Arg-Asp-Asp （RDD） receptor recognition site.

关 键 词：口蹄疫病毒 RGD受体结合位点 感染性cDNA克隆 病毒拯救 

分 类 号：S852.65[农业科学—基础兽医学]