大麦黄矮病毒外壳蛋白基因和运动蛋白基因酵母表达载体的构建

Yeast Expression Vectors′ Construction of Barley Yellow Dwarf Virus Coat Protein Gene and Movement Protein Gene

根据已知的大麦黄矮病毒GPV株系的外壳蛋白(Coat Protein CP)和移动蛋白(Movement Protein MP)基因序列合成了CP、MP基因的上下游引物,通过PCR扩增获得含有目的基因的片段,经过Sal I和Pst I双酶切、连接、转化、重组质粒的酶切鉴定及基因测序,构建了酵母表达载体pGBKT7-GPV-CP和pGBKT7-GPV-MP,用于在酵母双杂交分析中表达诱饵融合蛋白,为进一步筛选小麦cDNA文库内与大麦黄矮病毒互作的寄主因子、克隆寄主因子、推测其种类和功能奠定了基础。

The primers of CP and MP gene＇s upstream and downstream were synthesized according to the known BYDV-GPV strain＇s coat protein and movement protein gene sequence. Target genetic fragments included were aquired by means of PCR amplification. Through restrict endonuclase Sal Ⅰ / Pst Ⅰ, ligation, transformation, recombinant plasmids＇ restrict identification and gene sequencing, yeast expression vectors pGBKT7-GPV-CP and pGBKT7-GPV-MP were constructed,used for expressing the bait fusion protein in yeast two-hybrid analysis. The basis has been established to select the host factors which interact with the barley yellow dwarf virus in wheat cDNA library, clone the host factors and predict their species and functions.