利用酵母双杂交系统研究水稻条纹病毒三个功能蛋白的互作

Studies on the interactions between three major functional proteins of rice stripe virus using yeast two-hybrid system

通过RT—PCR扩增获得水稻条纹病毒（Rice stripe virus，RSV）的CP、SP和NSvc4基因，分别将其与酵母双杂交载体pGADT7和pGBK77相连，构建了重组子pGAD-CP、pGBK—CP、pGAD—SP、pGBK-SP、pGAD．NSnv4、pGBK-NSnv4。利用酵母双杂交系统研究了CP、SP、NSvc4三个蛋白的自激活、蛋白自身以及三个蛋白两两之间的互作情况。结果发现：RSV的CP、SP和NSvc4三个蛋白都不具有自激活现象；CP蛋白自身能够发生互作，SP和NSvc4蛋白未观察到自身互作现象；而CP、SP和NSvc4三个蛋白两两之间也未检测到互作现象的存在。该研究结果初步明确了RSV的CP、SP和NSvc4三个蛋白之间的关系，为追一步研究适三种蛋白的功能及RSV的致病机制奠定了基础。

Recently the series of researches focusing on the biology and molecular biology of RSV have been performing, but the relationship among CP, SP and NSvc4 remains poorly understood. Rice stripe virus （RSV） CP, SP and NSvc4 gene were amplified by RT-PCR. PCR products were cloned by pMD18-T and then linked with yeast two-hybrid vectors pGADT7 and pGBKT7, respectively, and recombinants pGAD-CP, pGBK-CP, pGAD-SP, pGBK-SP, pGAD-NSnv4 and pGBK-NSnv4 were constructed. By yeast two-hybrid, self-activation, self-interaction and interaction each other of CP, SP and NSvc4 were studied. Results demonstrated：that CP, SP and NSvc4 don＇t self-activate; CP can self-interact, while SP and NSvc4 cannot self-interact; Interaction each other of CP, SP and NSvc4 were also not detected. Based on these results, the relationship among CP, SP and NSvc4 were definitude primarily, and the results obtained the foundation for further research on function of virus proteins and pathogenetic mechanism of RSV.