小反刍兽疫病毒F基因真核载体的构建与表达  

Construction and expression of the eukaryotic expression vector with F gene of peste des petits ruminants virus

作　　者：朱学亮[1,2] 张强[1] 杨帆[1,2] 窦永喜[1] 骆学农[1] 岳城[2] 才学鹏[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甘肃省动物寄生虫病重点实验室,甘肃兰州730046 [2]新疆农业大学动物医学学院,新疆乌鲁木齐830052

出　　处：《中国兽医科学》2009年第5期405-409,共5页Chinese Veterinary Science

基　　金：国家农业公益性行业科研专项(200803018)

摘　　要：根据GenBank中小反刍兽疫病毒(Nigeria75/1株)F基因序列设计了引物,应用RT-PCR方法获得了目的基因,纯化回收后将其连接到克隆载体pMD18-T上。将鉴定与序列测定后的目的基因再克隆到真核表达载体pEGFP-N1上,经过鉴定,成功构建了含有小反刍兽疫病毒F基因的真核细胞表达载体pEGFP-N1-F。利用脂质体介导阳性pEGFP-N1-F质粒转染了BHK-21细胞。Western-blotting证实,表达的pEGFP-N1-F融合蛋白(90ku)具有免疫活性。According to the encoding sequence of the peste des petits ruminants virus（PPRV） F protein gene in GenBank,appropriate primers specific to F gene of PPRV were designed. A target DNA fragment was amplified by PCR and then the amplicon was cloned into pMD18-T vector. After identification by restriction digestion and sequencing, the specific DNA fragment was cloned into the eukaryotic expression vector pEGFP-N1. The plasmid was named pEGFP-N1-F after identification by restriction enzyme digestion and PCR amplification. The plasmid of pEGFP-N1-F was successfully constructed, and it was transfected into BHK-21 cells. The fusion protein from pEGFP-N1-F was 90 ku approximately and possessed immunological activity by Western-blot.

关 键 词：小反刍兽疫病毒 F基因 真核细胞表达载体 转染 免疫活性 

分 类 号：S852.659.5[农业科学—基础兽医学]