摘要
根据GenBank发表的传染性法氏囊病病毒 (IBDV) 5 2 70株VP2 基因序列 ,设计合成了 1对特异扩增IBDVVP2 基因的引物。以IBDV超强毒河南分离株HN0 1感染发病鸡法氏囊组织中提取病毒RNA来制模板 ,利用RT PCR扩增出了 1 .4kb的VP2 基因 ,将其克隆到pIREShyg载体上 ,构建了pIRES VP2 真核表达载体。然后通过磷酸钙共沉淀法转染CHO细胞 ,通过潮霉素筛选得到阳性克隆 ,间接免疫荧光实验 (IFA)鉴定VP2 在CHO细胞中的表达 ,并用RT PCR的方法从转录水平证实VP2 在CHO k1细胞中的表达 ,最终建立了CHO VP2
According to the published sequence of standard strain 52/70 of IBDV,a pair of primers was designed and synthesized,which can amplify vvIBDV's major protecting antigen VP 2 gene.A VP 2 gene's cDNA fragment of about 1.4kb was obtained through RT PCR method from HN01 strain of vvIBDV,VP 2 gene was amplified and cloned into pIREShyg Vector Eukarytic Vector pIRES VP 2was constructed.CHO cells coprecipitation with hygromycine selection,positive cells were screened.The protein VP 2 was expressed in the CHO cells proved by IFA.The VP 2 gene was transcript proved by RT PCR.So stable CHO/VP 2 cell strain was constructed.;
出处
《中国生物工程杂志》
CAS
CSCD
2004年第8期68-72,共5页
China Biotechnology
基金
国家"8 63"计划资助项目 ( 2 0 0 2AA2 45 0 5 1)
