摘要
通过对日本脑炎病毒(Japanese encephalitis virus,JEV)基因组中移码序列分析,设计合成1对引物,利用PCR扩增出NS1’移码片段,并将移码片段克隆入pET-32a表达载体中。重组质粒转化大肠杆菌BL21后,经IPTG于20℃过夜诱导,实现了目的片段的可溶性表达。对表达产物进行SDS-PAGE鉴定,可检测到分子质量约为25 ku的融合蛋白。以日本脑炎病毒野毒株和弱毒株感染小鼠血清为一抗,进行Western blot分析,原核表达蛋白仅与野毒感染小鼠血清发生特异性反应,说明该蛋白具有区分临床野毒感染和疫苗免疫的潜力。
The NS1＇ gene fragment resulted from ribosomal frameshifting in Japanese encephalitis virus（JEV） was amplified by RT-PCR and inserted into a prokaryotic expression vector pET-32a.The recombinant plasmid pET-32a-NS1＇ was transformed into Escherichia coli BL21 and induced to express by IPTG.The fusion protein with a molecular weight of 25ku was visible on the SDS-PAGE gel and purified with His-Bind Purification Kit.The Western blotting analysis showed that the NS1＇ protein could react with the polyclonal antibody from the mice immunized with wild strain of JEV,while the NS1＇ protein did not react with that from mice immunized with JEV vaccine.These results indicated that the NS1＇ protein had the potential to distinguish between wild strain infection and vaccine immunization in clinical practice.
出处
《畜牧与兽医》
北大核心
2013年第7期19-22,共4页
Animal Husbandry & Veterinary Medicine
基金
国家公益性行业(农业)科研专项(201203082)
关键词
日本脑炎病毒
NS1’蛋白
移码序列
原核表达
免疫反应性
Japanese encephalitis virus
NS1＇ protein
ribosomal frameshifting
prokaryotic expression
immunoreactivity
