南方、爪哇和花生根结线虫的快速灵敏的PCR鉴定方法

 为了研制南方、爪哇和花生根结线虫快速灵敏的检测和鉴定方法,分别分离了4个南方根结线虫和3个爪哇根结线虫特异性的随机扩增多态性DNA (RAPD)片段。在这些RAPD标记DNA序列的基础上,设计了多对SCAR PCR引物,并用源于国内外的南方、爪哇、花生、北方和象耳豆根结线虫群体验证其扩增特异性和灵敏度。最终确定了3对高效扩增的SCAR引物,它们组合使用可以可靠灵敏地鉴定南方、爪哇和花生根结线虫。3对引物的扩增灵敏度达1/3条的二龄幼虫、雄虫或雌虫,这表明本研究研制的PCR鉴定法可用于生产实践中土样和根样中3种根结线虫快速灵敏的鉴定。

PCR Assays for Rapid and Sensitive Identification of Three Major Root-Knot Nematodes,Meloidogyne incognita,M.javanica and M.arenaria

To develop a rapid and sensitive method for the detection and identification of Meloidogyne incognita, M. javanica and M. arenaria, four and three random amplified polymorphic DNA (RAPD) fragments specific for M. incognita and M. javanica, respectively, were identified. Based on the sequences of these RAPD markers, various sequence characterized amplified region (SCAR) primers were designed and tested for their amplification specificity and efficiency against populations of M. incognita, M. javanica, M. arenaria, M. hapla and M. enterolobii. This resulted in three pairs of SCAR primers that were used in combination to reliably and sensitively identify M. incognita, M. javanica and M. arenaria. The SCAR markers can be readily amplified from one third of a single second-stage juvenile, male or female, thus demonstrating that the SCAR-based PCR assays have practical value for routine identification of M. incognita, M. javanica and M. arenaria in soil and root samples.