葡萄卷叶病毒的纯化、血清学研究及其在脱毒组培苗检测中的应用  

PURIFICATION, SEROLOGY AND DETECTION OF GRAPEVINE LEAF ROLL VIRUS

作　　者：蔡文启[1] 徐绍华[1] 莽克强[1] 孟保中[2] 马云霞[2] 

机构地区：[1]中国科学院微生物研究所,北京100080 [2]河北省农林科学院昌黎果树研究所,昌黎066600

出　　处：《微生物学报》1997年第5期385-392,共8页Acta Microbiologica Sinica

基　　金：国家"八五"攻关资助项目

摘　　要：将具有典型葡萄卷叶病(Grapevine leafroll diseas,GLRD)症状的葡萄组织,经差速和硫酸铯—蔗糖密度梯度离心,提纯了GLRV,并制备了兔抗血清。电镜下可观察到长度从600～2000nm的线形病毒颗粒,其中以1400nm左右为主。免疫电镜结果表明线形病毒颗粒能被美国的NY-1分离株抗血清(Ⅲ型)所修饰。在间接ELISA中提纯制品与GLRV的Ⅲ、Ⅳ、Ⅱ型抗血清均能产生免疫反应。与Ⅲ型抗血清产生较强的免疫反应,Ⅳ型次之,Ⅱ型最弱。在SDS-免疫双扩散实验中病组织韧皮部粗提液与GLRV的Ⅲ,Ⅳ、Ⅱ型抗血清均产生免疫沉淀线。从而推测我国葡萄园内的葡萄卷叶病很可能由2种或3种卷叶病毒感染所致.采用A蛋白夹心酶联免疫吸附试验(PAS-ELISA)检测葡萄试管苗,Ⅲ型抗血清和自制抗血清的平行测试结果基本相符,共获得11个生食葡萄和10个山葡萄品种的脱葡萄卷叶病毒和扇叶病毒的组培苗,扩繁后田间试种表现出良好的农艺性状。Closterovirus-like particles associated with grapevine leafroll disease were purified from stem phloem tissue by differential centrifugation and cesium sulphate-sucrose density gradient centrifugation. The particles were between 660-2000nm long. Most of them were about 1400nm long and also decorated by the antiserum against GLRV NY-1 isolate. The purified preparation was tested for their serological relatedness in indirect ELISA. The results indicated that these closterovirus-like particles reacted with serotype Ⅱ, Ⅲ and Ⅳ of GLRV. The antiserum to NY-1 (type Ⅲ) reacted more strongly than type Ⅳ (to VA-4) and type Ⅱ. (to 1800nm). In sodium dodecyl sulfate immunodiffusion test, the extract of diseased petiole reacted with serotypeⅢ, Ⅳ and Ⅱ. It is possible to be infected by 2 or 3 closterovirus-like particles in China. We have made antiserum to the purfied closterovivus-like GLRV and set up a PAS-ELISA to detect GLRV-free grapevine plantelets. We have obtaimed 21 varieties of GLRV-free and GFLV-free grapevine. They shown high quality and quantity in field test

关 键 词：植物病毒 葡萄卷叶病毒 纯化 血清学 检测 

分 类 号：S432.41[农业科学—植物病理学] S436.631.1[农业科学—农业昆虫与害虫防治]