贝西滑刃线虫双重PCR检测方法研究  

Development of duplex PCR assay for detection of Aphelenchoides besseyi

作　　者：林宇[1] 王金成[1] 张宇含 董立[2] 顾建锋[3] 李志勇[2] 

机构地区：[1]天津出入境检验检疫局,天津300456 [2]河北省农林科学院谷子研究所,河北石家庄050035 [3]宁波出入境检验检疫局,浙江宁波315012

出　　处：《河北农业大学学报》2016年第1期80-83,93,共5页Journal of Hebei Agricultural University

基　　金：国家质检总局课题(2011IK176);国家科技支撑计划项目(2012BAK11B03);谷子产业技术体系(CARS-07-05-B)

摘　　要：为实现贝西滑刃线虫的高效、准确鉴定,根据贝西滑刃线虫核糖体28SrRNA-D2/D3片段的保守区域设计了特异性引物,并结合植物寄生线虫通用引物作为内标,构建了贝西滑刃线虫的一步双重PCR检测体系。结果表明:特异性引物GU-F/GU-R对贝西滑刃线虫具有高度的特异性,扩增出245bp的特异性目标片段,有效检测灵敏度可达0.125ng/μL线虫DNA模板量。该方法特异性好、灵敏度高、稳定性强,适用于贝西滑刃线虫的快速检测。To detect Aphelenchoides besseyi accurately and effectively, specific primers were designed based on the conserved regions of 28S rRNA-D2/D3 in the ribosome, and combined with the universal primers for the amplification of ribosomal ITS region in plant-parasitic nem-atodes. The duplex PCR method for detecting A. besseyi was therefore established. The results showed that a fragment of 245 bp was amplified from A. besseyi by specific primers GU- F/GU-R. The sensitivity test showed that the duplex PCR can detect 0. 125 ng/μL of nema- tode DNA. The results indicated the duplex PCR method had advantages of high specificity, sensitivity and stability. It can be effectively used in the detection of A. besseyi.

关 键 词：贝西滑刃线虫 双重PCR 特异性引物 分子鉴定 

分 类 号：S476[农业科学—农业昆虫与害虫防治]