摘要
应用RT-PCR技术扩增出犬瘟热病毒(CDV)核衣壳(N)蛋白基因的高度保守序列,将其克隆至质粒pMD18-T中,获得了重组质粒pMD18-T-N。将N基因的目的片段克隆到表达载体pGEX-4T-1中谷胱甘肽转移酶(GST)基因的下游,并将该重组质粒转化大肠杆菌BL21株,经IPTG诱导,N基因融合蛋白获得了高效表达。SDS-PAGE电泳和Western-blot分析结果显示,表达产物的分子质量为55 ku,与CDV标准阳性血清呈阳性反应。表明,大肠杆菌表达的CDV N蛋白在免疫原性上与天然N蛋白具有一定的相似性,可作为诊断用抗原。
A highly conserved sequence of nucleocapsid protein gene of canine distemper virus（CDV） was amplified by RT-PCR, then the product was cloned into pMD18-T vector to construct recombinant plasmid pMD18-T-N and sequenced. The gene was inserted into the downstream of GST of pGEX-4T-1 vector. The plasmid pGEX-4T-1-N was transformed into Escherichia coli BL21 for expression under induction with IPTG. SDS-PAGE analysis showed that the molecular weight of the expressed product was 55 ku. Western-blot analysis showed that the recombinant protein had a positive reaction with standard positive serum against CDV. The results indicated that the expressed N protein was similar to the natural nucleocapsid protein on immunogenicity,which could be used as antigen for diagnosis.
出处
《中国兽医科学》
CAS
CSCD
北大核心
2007年第5期378-381,共4页
Chinese Veterinary Science
关键词
犬瘟热病毒
核衣壳蛋白
免疫原性
canine distemper virus
nucleocapsid protein
immunogenicity
