鸭坦布苏病毒E蛋白主要抗原区域的原核表达与鉴定

Prokaryotic Expression and Identification of Duck Tembusu Virus Main Antigenic Region of E Protein

根据GenBank中发表的鸭坦布苏病毒(DTMUV)基因序列,设计合成一对特异性引物,RT-PCR扩增966bp E基因片段并克隆至pMD18-T载体中,经鉴定正确后,将其定向克隆至pET30a表达载体中,经鉴定正确后,重组阳性质粒转化大肠埃希菌BL21(DE3)表达菌,经IPTG诱导获得了以包涵体形式表达的重组蛋白。采用镍离子亲和层析柱对重组蛋白进行纯化,Western blot检测证明该重组蛋白可与抗血清发生反应。

Based on published duck Tembusu virus（DTMUV）gene sequences in GenBank,apair of specific primers were designed,966bp E gene fragment was amplified by RT-PCR and cloned into pMD18-T vector,the fragment was cloned into expression vector pET30acorrectly after identification,and then the positive plasmid transformed to BL21expression bacteria.After induction by IPTG,the recombinant proteins were expressed in inclusion body forms.Using nickel ion affinity chromatography,the recombinant proteins were purified.Western blot showed the recombinant protein had good antigenicity,which laid a foundation for E protein structure and function research,and research for detection kit of DTMUV antibody and new type vaccines.