利用酵母双杂交系统筛选BmNPV PE38的互作蛋白

利用酵母双杂交技术,以家蚕核型多角体病毒(BmNPV)的PE38蛋白为诱饵,从感染BmNPV的家蚕中肠组织cDNA文库中筛选与之相互作用的宿主蛋白编码基因,为探讨BmNPV PE38在侵染家蚕过程中的作用提供线索。将成功构建的酵母诱饵表达载体pGBKT7-Pe38转化到酵母菌Y2HGold中,并将含有感染BmNPV的家蚕中肠组织cDNA文库的Y187酵母细胞与含有pGBKT7-Pe38的酵母细胞共同培养,初筛出8个有匹配结果的候选蛋白基因,经功能搜索后筛选到2个与病毒复制有关的蛋白基因,通过测序和生物信息学分析,这2个基因编码的蛋白质可能是与BmNPV的PE38蛋白有相互作用的候选蛋白,分别为家蚕丝氨酸蛋白酶和过氧化氢酶。

Identification of BmNPV PE38-Interactive Proteins Using Yeast Two-hybrid System

In order to provide clues for exploring the role of PE38 protein of Bombyx mori nucleopolyhedrovirus(Bm-NPV) in its infection to Bombyx mori,PE38 protein of BmNPV was used as bait in yeast two-hybrid system to identify genes encoding host proteins interactive with PE38 in a cDNA library constructed from BmNPV infected Bombyx mori midgut.The successfully constructed yeast bait expression vector pGBKT7-Pe38 was transformed into yeast strain Y2HGold.Then,Y187 yeast cells harboring cDNA library constructed from BmNPV infected silkworm midgut were co-cultured with yeast cells containing pGBKT7-Pe38.As a result,8 candidate protein coding genes were found to have hits in GenBank database.Functional searches led us to identify 2 protein genes related to virus replication.Sequencing and bioinformatic analysis revealed that these two genes encoded proteins,Bombyx mori serine protease and catalase,were candidate proteins interactive with BmNPV PE38 protein.