传染性法氏囊病病毒VPP5酵母双杂交诱饵载体的构建及互作宿主细胞蛋白的筛选

Construction of the Bait Vector of Infectious Bursal Disease Virus VP5 and Screening the VP5 Interacted Proteins of Host Cell using Yeast Two-hybrid System

VP5蛋白是传染性法氏囊病病毒(Infectious bursal disease virus,IBDV)编码的一种非结构蛋白,在病毒感染、释放、致病性方面起着重要作用。为了研究VP5蛋白与宿主细胞的相互作用,本试验首先将IBDV VP5基因克隆入p GBKT7,构建诱饵载体p GBGt VP5,通过自激活试验证明其没有自激活活性,对酵母细胞没有毒性。然后,运用酵母双杂交技术,从鸡胚成纤维细胞(CEF)c DNA文库中筛选与VP5互作蛋白,经初筛、测序和回转验证,获得5个互作宿主细胞蛋白,为进一步深入研究IBDV VP5与宿主互作的效应和机制奠定基础。

Viral protein 5 （VPS）,a non-structural plays an important role in viral infection, release and protein encoded by infectious bursal disease virus （IBDV）genome, pathogenicity. To study the interaction between VP5 and the hostcell, VP5 full-length cDNA was amplified and inserted into pGBKT7 vector to construct a bait vector （pGBGtVPS）. The results of autoactivation assay showed that pGBGtVP5 vector had no autoactivation and virulence in Y2H Gold yeast cell. After screening,sequencing,and confirming interaction,5 kinds of interacted proteins were obtained from the expression library of chicken embryo fibroblasts （CEF） cell using the Matchmaker Gold Yeast Two-Hybrid System. This study laid the foundation for further researches on the effect and mechanism of the interaction between IBDV VP5 and its host cell.