实时荧光PCR检测马铃薯白线虫技术研究.pdfVIP

2009 年第 4期 植物检疫 PLANT QUARANT INE V ol. 23 No. 4 PCR 1 1 1 2 葛建军 曹爱新 陈洪俊 M ariceMoens ( 1 100029 2 M erelbeke 9820) 1 1 1 Real tim e PCR ofG lobod era p a llida w ith TaqM an probe. Ge Jianjun , CaoA ixin , Chen H ongjun , M arice 2 M oens ( 1 Institute of An mi al and Plant Quarantine, Ch inese A cademy of Inspection and Quarantine, Beijing 100029, Ch ina, 2 Belgium A griculturalReearch Centre, M erelbeke, 9820, Belgium ) Abstract Potato cyst nematodes are one of themost mi portant plant pests infested potatoes, also are p lant quar antine nematodes concerned specially by Ch inese Governm ent. To carry out rap id and reliable species detection method ofG lobodera p allida, w e designed prmi ers and TaqM an probe depended on sequence of ITS. The prmi ers and probe w ere evaluated on 1 samp les of PCN popu lations and 4 other cyst nem atodes. The PCR assay for spe cies detection is h ighly sensitive, w h ich could detect the template concentration as low as 10 fg. Th ism ethod can identify and quantify single air- dried cyst and juven ile of G. p allida, and also allow s the detection of species m ixtures in a sample. Furthermore, the realtmi e PCR assay w ithTaqM an provides a rap idmeans for smi ultaneous detection and quantification of PCN and is therefore particu larly useful for routine d iagnoses. K eywords G lobodera p a llida, molecu lar detection, Real tmi e PCR 马铃薯胞囊线虫是马铃薯上最重要的有害生物之 一, 也是我国 别关注的重要植物检疫性线 虫针对马铃薯白线虫 ITS序列, 我们设计了引物和 TaqM an探针, 使用 1 种马铃薯胞囊线虫群体和 4种 其它胞囊线虫样品进行验证, 可高度灵敏地检测单 个马铃薯白线虫的胞囊或幼虫, 最高检测灵敏度达到 10fg; 同时开展了混合样品和未知样品的检测, 证明了引物的专化性和TaqM an 探针 异性该检测方法可 自动化检测马铃薯白线虫并进行定量, 适合进行标准化的常规检测 马铃薯白线虫 分子检测 实时荧光 PCR Q9 9. 1