表达水泡性口炎病毒G蛋白的重组腺病毒的构建与鉴定  

Construction and identification of recombinant adenovirus expressing vesicular stomatitis virus G protein

作　　者：薛晓娟[1] 金红岩[1] 隋修锟[1] 李文超[1] 李刚[1] 

机构地区：[1]中国农业科学院北京畜牧兽医研究所动物营养学国家重点实验室/农业部兽用药物与兽医生物技术北京科学观测实验站,北京100193

出　　处：《中国兽医科学》2015年第5期448-452,共5页Chinese Veterinary Science

基　　金：国家科技支撑计划项目(2013BAD12B05);国家自然科学基金项目(31172342);转基因重大专项(2014ZX0801203B);国际合作项目(D32025/17453);WSPA公益项目(2013-hx-04);创新团队基金项目(ASTIP-IAS15)

摘　　要：为了以复制缺陷型人5型腺病毒为表达载体,构建表达水泡性口炎病毒印第安纳型(VSV-IN)G蛋白的重组腺病毒,利用PCR扩增编码VSV-IN G蛋白的基因,并将其克隆至腺病毒穿梭载体pacAd-CMV K-NpA中;再将含有目的基因的重组穿梭质粒与腺病毒骨架载体质粒用PacⅠ酶切线性化后共转染AAV-293细胞,获得重组腺病毒rAd-VSV-IN-G。该重组腺病毒于AAV-293细胞连续传代至第20代效价稳定。间接免疫荧光试验和Western-blot检测结果显示,G蛋白在重组腺病毒中可以被正确表达。上述研究成果为VSV重组腺病毒疫苗的研制奠定了基础。In order to construct the recombinant adenovirus vectors expressing vesicular stomatitis virus Indiana type(VSV-IN)G protein using the human adenovirus expression vector,the G gene of VSVIN was amplified by PCR.The gene was cloned into the adenovirus shuttle vector pacAd-CMV K-NpA.The recombinant shuttle plasmid and adenovirus backbone plasmid was linearized by digestion with PacⅠ,and transfected into AAV-293 cells to obtain recombinant adenovirus rAd-VSV-IN-G.The recombinant adenovirus in AAV-293 cells were serially propagated to 20 generations with high potency and stability.By indirect immunofluorescence assay and Western-blot detection,the G protein was expressed in the recombinant adenovirus.The above-mentioned results showed that the adenovirus expressing VSV G protein can be furtherly used to develope vaccines against VSV.

关 键 词：水泡性口炎病毒 G蛋白 重组腺病毒 

分 类 号：S852.65[农业科学—基础兽医学]