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.
出处
《淡水渔业》
CSCD
北大核心
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)
