免疫磁珠RT-PCR法检测葡萄扇叶病毒  

Detection of Grapevine fanleaf virus by immunomagnetic separation and RT-PCR

作　　者：魏梅生[1] 尤佳 马洁[1] 李桂芬[1] 张永江[1] 

机构地区：[1]中国检验检疫科学研究院植物检疫研究所,北京100029 [2]甘肃出入境检验检疫局

出　　处：《植物检疫》2014年第4期32-35,共4页Plant Quarantine

基　　金：国家质量监督检验检疫总局课题(2012IK302)

摘　　要：葡萄扇叶病毒引起的葡萄扇叶病是葡萄生产上的主要病害之一,建立简便、灵敏、准确的检测方法是葡萄健康生产的基础。将葡萄扇叶病毒抗体和0.5μm大小的超顺磁性磁珠SiMAG-PGL偶联,制备出特异性免疫磁珠,用于分离和富集检测样品中的病毒。富集的病毒可直接用于RT-PCR检测,本文所建立的检测方法可有效地检测到葡萄扇叶病毒的3个分离物,而检测烟草环斑病毒、南芥菜花叶病毒的结果为阴性,检测的特异性较强。此法检测提纯的葡萄扇叶病毒灵敏度可到0.01 ng/mL,检测葡萄扇叶病毒感染的苋色藜叶片可稀释到10-6,该方法适用于植株体内低浓度病毒的检测。Grapevine fanleaf virus （GFLV）, the causal agent of grapevine fanleaf disease, is one of the most damaging virus affecting grapevine. Establishment of vineyards free of GFLV is based on efficiently testing method. A rabbit polyclonal antibody against GFLV was coupled with 0.5μm SiMAG-PGL magnetic bead. The immunomagnetic bead could be used to separate and trap the virus from the testing sample. The concentrated virus was tested by RT-PCR. The method could detect 3 isolates of GFLV. There was no cross reaction with Tobacco ringspot virus and Arabis mosaic virus. The sensitivity of the method was 0.01 ng/mL for detecting purified GFLV. Extract from GFLV infected Chenopodium amaranticolor leaf was detected to a dilution of 10^-6. It is recommended to use this method for detecting low concentration of virus in plant.

关 键 词：葡萄扇叶病毒 免疫磁珠 RT-PCR 

分 类 号：S436.631.1[农业科学—农业昆虫与害虫防治]