抑制南方水稻黑条矮缩病毒非结构蛋白P5-1的表达阻碍病毒在昆虫体内的增殖  

Knockdown of nonstructural protein P5-1 of Southern rice black-streaked dwarf virus prevents viral multiplication in vector insects

作　　者：曹金强[1] 王针针 陈红燕[1] 毛倩卓[1] 贾东升[1] 魏太云[1] 

机构地区：[1]福建农林大学植物病毒研究所福建省植物病毒学重点实验室,福州350002

出　　处：《植物保护学报》2015年第3期418-424,共7页Journal of Plant Protection

基　　金：国家自然科学基金(31130044;31400136;31401712)

摘　　要：为明确南方水稻黑条矮缩病毒(Southern rice black-streaked dwarf virus,SRBSDV)编码的非结构蛋白P5-1参于SRBSDV在介体白背飞虱体内侵染过程中的作用机制,通过原核表达蛋白制备SRBSDV编码的非结构蛋白P5-1的多克隆抗体,并应用Western blot和免疫荧光标记法检测抗体的特异性,以注射法将来源于P5-1基因的dsRNA(ds P5-1)注入获毒1 d的白背飞虱体内,5 d后通过免疫荧光标记法检测ds P5-1对SRBSDV在白背飞虱体内增殖的影响,同时以注射来源于GFP基因的dsRNA(ds GFP)为对照。结果显示,Western blot和免疫荧光标记分别检测到SRBSDV侵染水稻和白背飞虱表达的P5-1蛋白,表明所制备的P5-1抗体具有特异性。ds GFP处理的对照组白背飞虱带毒率高达81%,而ds P5-1处理的白背飞虱带毒率仅为21%,且P5-1蛋白的表达和SRBSDV在昆虫体内的增殖均受到抑制。表明P5-1蛋白是病毒在昆虫体内增殖的关键因子,可作为阻断病毒在昆虫体内增殖的理想靶标。To investigate the function of the nonstructural protein P5-1 of Southern rice black streaked dwarf virus (SRBSDV) in the insect vector white-backed planthopper (WBPH),Sogatella furcifera (Horvdth),here the prokaryotic expression of P5-1 was performed,followed by the preparation of polyclonal antibody against P5-1.Then the specificity of polyclonal antibody was tested by western blot and immunofluorescence.WBPH was microinjected with dsRNA targeting P5-1 gene (dsP5-1) of SRBSDV or dsRNA targeting GFP gene (dsGFP) at day one after virus acquisition.Five days later,the effect of dsP5-1 on viral mulitiplication in the insect vector was examined using immunofluorescence assay.The results showed that the protein P5-1 was detected in infected rice plants using western blot,and the specificity of polyclonal antibody against P5-1 was confirmed in viruliferous WBPH by using immunofluorescence.The rate of viruliferous insects in the control was 81％,while that of dsP5-1 microinjection was 21％.dsP5-1 significantly inhibited the expression of P5-1 and multiplication of SRBSDV in the body of WBPH vector.These results suggested that P5-1 was the key factor influencing viral multiplication in insect vector,which may serve as a target for blocking viral multiplication in WBPH.

关 键 词：南方水稻黑条矮缩病毒 P5-1蛋白 抗体制备 RNA干扰 增殖 

分 类 号：S435.111.4[农业科学—农业昆虫与害虫防治]