草鱼呼肠孤病毒VP6蛋白基因植物表达载体的构建  

Construction of plant-based expression vector of grass carp reovirus VP6 gene

作　　者：周勇[1,2] 范玉顶[2] 徐进[2] 李艳秋[2] 肖艺[1] 曾令兵[1,2] 

机构地区：[1]华中农业大学水产学院,武汉430070 [2]中国水产科学研究院长江水产研究所,湖北荆州434000

出　　处：《淡水渔业》2009年第4期40-44,共5页Freshwater Fisheries

基　　金：农业公益性行业科研专项(200803013)

摘　　要：以据草鱼呼肠孤病毒(GCRV)VP6蛋白的全基因序列(GeneBank AF403394)为模板,采用RT-PCR构建了草鱼呼肠孤病毒VP6蛋白基因植物表达载体的构建。结果显示:实验构建的pCR 2.1-VP6重组质粒含有NcoⅠ和BglⅡ酶切位点;pCR 2.1-VP6重组质粒经PCR扩增和测序显示含有1.3 Kbp的草鱼呼肠孤病毒VP6基因读码框片段。将目的片段VP6酶切、克隆到携带有绿色荧光蛋白(GFP)基因的植物表达载体pCAMBIA1302中,再经酶切、PCR扩增和序列测定显示,pCAMBIA1302-VP6含有1.3 Kbp的草鱼呼肠孤病毒VP6基因片段,说明已插入植物表达载体pCAMBIA1302绿色荧光蛋白基因前,成功构建了融合表达草鱼呼肠孤病毒VP6蛋白和绿色荧光蛋白的植物表达载体pCAMBIA1302-VP6。The coding region of grass carp reovirus（GCRV） VP6 gene was amplified by RT-PCR with primers based on the gene sequence in GeneBank（AF403394） from the viral RNA genome extracted in virus infected Ctenopharyngodon idellus Kidney（CIK） cells.1.3 kb PCR product was ligated to pCR 2.1 vector,and after identified with enzyme digestion,PCR detection and sequencing,then the 1.3 kb cording sequence of GCRV VP6 gene was cloned into the plant-based expression vector pCAMBIA1302which had a green fluorescent protein（GFP） gene insert.Thus,a fusion-expression vector of grass carp reovirus VP6 gene and GFP gene was constructed.This study has established a foundation for preparing plant-based edible vaccine against the grass carp hemorrhage.

关 键 词：草鱼呼肠孤病毒 VP6基因 植物表达载体 构建 

分 类 号：S941.41[农业科学—水产养殖] Q782[农业科学—水产科学]