肠道病毒71型重组VP2蛋白原核表达及初步鉴定  

Prokaryotic expression of recombinant VP2 protein of Enterovirus 71 and its preliminary identification

作　　者：施静[1] 朱冰[1] 钟家禹[1] 王长兵[1] 许甜甜[1] 

机构地区：[1]广州医学院附属广州市妇女儿童医疗中心实验室,广东广州510120

出　　处：《国际检验医学杂志》2013年第12期1494-1496,共3页International Journal of Laboratory Medicine

基　　金：国家自然科学基金资助项目(30972630)

摘　　要：目的利用大肠杆菌表达系统表达并纯化肠道病毒71型(EV71)VP2蛋白,并对重组蛋白功能进行初步鉴定。方法 PCR扩增EV71 VP2基因,在测序正确的基础上,将其插入表达载体形成pET32a(+)/VP2,转化BL21(DE3),IPTG诱导表达,经十二烷基硫酸钠聚丙烯酰胺凝胶电泳(SDS-PAGE)初步分析蛋白表达产物,利用Ni+亲和层析柱对重组蛋白进行纯化,经Western blot初步验证其抗原性。结果成功构建pET32a(+)/VP2重组质粒,经大肠杆菌表达,纯化获得相对分子质量约为48000的融合蛋白,经Western blot证实其抗原性良好。结论表达并鉴定了VP2抗原,为EV71疫苗研制及检测试剂盒的研发奠定了基础。Objective To expression and purify VP2 protein of Enterovirus 71（EV71） through E.coli expression system and identify the recombinant protein function preliminarily.Methods EV71 on the basis of correct sequencing.BL21（DE3） was transformed and IPTG expression VP2 gene was amplify by PCR and then inserted into expression vector to form pET32a（＋）/VP2 induced.Sodium dodecyl sulfate polyacrylamide gel electrophoresis（SDS-PAGE） was used to preliminarily analyze the protein expression product,Ni＋ affinity columns were adopted to purified the recombinant protein and Western blot was employed to preliminarily validate its antigenicity.Results pET32a（＋）/VP2 recombinant plasmid was successfully constructed.Purified fusion protein of relative molecular mass 48 000 was obtained through expression in E.coli and demonstrated good antigenicity by Western blot identification.Conclusion Expression and identification of VP2 antigen in the study provide a foundation for research and manufacture of EV71 vaccine and development of test kits.

关 键 词：肠道病毒71型 VP2蛋白 原核表达 

分 类 号：R392[医药卫生—免疫学]