AsiaⅠ型口蹄疫病毒VP1基因结构预测及亚细胞定位  

Structure prediction and subcellular localization of VP1 protein from foot-and-mouth disease virus serotype Asia

作　　者：张克山[1] 尚佑军[1] 郭建宏[1] 郑海学[1] 田宏[1] 靳野[1] 刘永杰[1] 刘湘涛[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,甘肃兰州730046

出　　处：《中国兽医学报》2012年第11期1669-1673,共5页Chinese Journal of Veterinary Science

基　　金：国家现代肉羊产业技术体系资助项目(nycytx-39);农业微生物学国家重点实验室开放课题资助项目(AMLKF201005)

摘　　要：利用PCR方法扩增到VP1基因,序列测定后利用Mega4.0、swiss-model、GOR4和RasMol软件预测了VP1基因的二、三级结构。将VP1基因融合EGFP基因后定向克隆入PcDNA3.1(+)真核表达载体,构建正确的重组质粒命名为PVP1E,在脂质体介导下将PVP1E质粒转染BHK-21细胞,Western Blotting试验证实VP1基因成功表达,经DAPI细胞核染色后在共聚焦显微镜下观察VP1亚细胞定位。结果表明本研究预测了VP1基因的二、三级结构,其在BHK-21细胞中呈现以细胞核为主的弥散性分布,VP1基因亚细胞定位及结构预测为进一步深入探究AsiaⅠ型FMDV VP1结构和功能提供丰富的资料。VP1 gene was amplified and sequenced, Softwares of Mega4.0, swiss-model, GOR4 and RasMol were used to predict secondary and 3-D structures for VP1 gene. Then VP1 gene was sub- cloned into eukaryotic expression vector PcDNA3. 1（＋） directionally after fused with EGFP（en- hanced green fluorescent protein） , the recombinant plasmid named PVPIE was identified success- fully via restriction enzyme digestion analysis, DNA sequencing. The PVP1E was transfeeted into BHK-21 cells mediated by Liposomes 2000. Expression of VP1 gene in BHK-21 cells was con- firmed by western-blot. Subcellular localization of VP1 gene was observed by eonfocal microscopy after DAPI nuclear staining. The results indicated that VP1 gene structures were successfully pre- dicted,VP1 protein displayed diffuse distribution and located mainly in nucleus of BHK-21 cells. This research accumulates useful materials and provids powerful analysis tools for further stud- ying structure and function of VP1 gene.

关 键 词：AsiaⅠ型FMDV VP1基因 结构预测 亚细胞定位 

分 类 号：S852.65[农业科学—基础兽医学]