锯缘青蟹(Scylla serrata)呼肠孤病毒5个结构蛋白互作分析

摘要

采用酵母双杂交系统,对青蟹呼肠孤病毒(Scylla serrata reovirus)5个结构蛋白(VP3,VP6,VP9,VP11,VP12)间的双向互作进行了分析.将5个结构蛋白对应的ORF分别亚克隆至诱饵载体pGBKT7和猎物载体pGADT7,成功构建了10个酵母双杂交重组载体.将重组诱饵载体或重组捕获载体转化至酵母菌Y2H Gold中进行自激活检测,结果显示5个结构蛋白均不能激活酵母菌报告基因HIS3的表达,表明5个病毒蛋白均不具有自激活活性.通过酵母双杂交实验,从5个结构蛋白25个可能性互作对中,共筛选出2个双向互作对(VP 11 &VP6,VP11&VP12)和3个自身互作对(VP3&VP3,VP11&VP11,VP12&VP12).本研究是有关SsRV结构蛋白互作的首次报道.%Each pair-wise combination among five structural proteins of Scylla serrata reovirus (SsRV) was analyzed systematically for possible interactions.Potential binary protein interactions among individual structural proteins of SsRV were screened by yeast two-hybrid (Y2H).All the five structural genes (VP3,VP6,VP9,VP11 and VP12) were tested pair-wise against each other in duplicate.None of the proteins was found to autoactivate the HIS3 reporter gene when expressed in the yeast.Of 25 combinations among the five VPs that accounted for two binary and three self associations were identified to be stable and functional within the yeast.The two binary interactions comprised VP11&VP6 and VP11 &VP12,while the self association comprised VP3&VP3,VP11 &VP11,and VP12&VP12.None of these interactions have been documented previously for SsRV.