半巢式RT-Realtime PCR检测番茄丛矮病毒  

Dectection of Tomato Bushy Stunt Virus by Semi-nested RT-Realtime PCR

作　　者：粟智平 杨益娥 赵伟铎 李金庆 邓丛良[3] 尹伟力 

机构地区：[1]烟台出入境检验检疫局,山东烟台264000 [2]湖南省会同县农业局 [3]北京出入境检验检疫局

出　　处：《现代农业科技》2015年第18期148-150,共3页Modern Agricultural Science and Technology

基　　金：烟台市科学技术发展计划项目(2008324);国家973研课题(2011CB932800);山东检验检疫局科研项目(SK201305);山东检验检疫局科研项目(SK2011)

摘　　要：番茄丛矮病毒(ToBSV)是我国进境植物检疫对象,也是进境种子苗木等必检项目。该病毒能侵染番茄、曼陀罗、葡萄、豇豆等20余科120余种植物。该研究根据ToBSV全基因组中p33蛋白基因,设计引物和Taq Man探针,建立了半巢式实时PCR检测ToBSV的新方法。该研究巧妙地融合了巢式PCR和Taq Man探针技术,第二步半巢式-RT-Realtime PCR既进一步放大了第一步检测信号,也是对第一步PCR产物的确认,与单一的巢式PCR或Realtime PCR等方法相比其检测的准确性、灵敏度更高。该方法检测灵敏度可达50 fg/μL植物总RNA。Tomato bushy stunt virus（ToBSV）is a plant quarantine object in China ,and it is a required inspection item for the imported grapes and so on.This virus can infect 120 kind of 20 families of plants,such as tomato,Datura stramonium L. ,grape and cowpea.Three nested primers and one TaqMan probe were designed according to the p33 protein gene of the ToBSV ,with which we set up a new method of semi-nested RT-Realtime PCR to detect the ToBSV.The nest-PCR and the probe-detection technique were combined maneuverably in this study.The semi-nested RT-Realtime PCR in the second step can both magnify the sigal and validate the result of the first step ,which can provide a more sensitive and specific detection of the ToBSV.The sensitivitv of the method can reach 50 fg/μL of total plant RNA.

关 键 词：番茄丛矮病毒(ToBSV) 半巢式RT—RealtimePCR 检测 

分 类 号：S41-30[农业科学—植物保护]