大豆霜霉病菌rDNA ITS区的分子探针的设计与应用  

Development and Application of Molecular Probe of Soybean Downy Mildew Based on Peronospora manshurica(Naoun.)Sydow rDNA ITS

作　　者：刘艳[1] 沙爱华[1] 陈海峰[1] 周蓉[1] 巴红平[1] 陈水莲[1] 杨中路[1] 邱德珍[1] 吴学军[1] 单志慧[1] 周新安[1] 

机构地区：[1]中国农业科学院油料作物研究所,农业部油料作物生物学重点开放实验室,湖北武汉430062

出　　处：《华北农学报》2012年第2期230-233,共4页Acta Agriculturae Boreali-Sinica

基　　金：农业产业技术体系(nycytx-004);"十一五"国家科技支撑计划(2009-BADA8B02);作物种质资源保护项目课题(NB2010-2130135-25-14-18)

摘　　要：大豆霜霉病菌是引起大豆病害的重要病原之一,采用真菌18～28 S间的内转录间隔区(Internal transcribed spacer,ITS)通用引物ITS1和ITS4扩增大豆霜霉病菌和其他外群真菌的基因组DNA,扩增出约500 bp的片段;通过克隆测序大豆霜霉病菌的ITS全序列并与GenBank中霜霉菌属其他种的ITS序列比对,设计出大豆霜霉病菌的特异性引物PM1和PM2。用此特异引物可以从大豆霜霉菌株中扩增出380 bp的特异性片段,而其余9个参试菌株和大豆组织的PCR反应结果为阴性,灵敏度试验证明,可以检测到目标DNA的浓度为0.1 pg。该方法可用于快速、准确和灵敏地检测大豆霜霉病菌,为快速监测组织中霜霉病菌潜伏侵染并及早采取防治措施提供积极的指导作用。Peronospora manshurica （Naoun.）Sydow is one of the important pathogen causing soybean diseases. A approximately 500 bp fragment was amplified via universal fungal primers ITS1 and ITS4 derived from internal transcribed spacer region between the 18S - 28S in soybean downy mildew Peronospora manshurica （ Naoun. ） Sydow. Specific primer pairs PM1 and PM2 was designed after homological analysis of rDNA ITS sequences among Peronospora manshurica（ Naoun. ）Sydow and other species based on soybean downy mildew by cloning and sequen- cing of ITS sequences in GenBank. A 380bp fragment was amplified with this specific primer from soybean downy mildew strains and infected leaves,while no PCR products was amplified from other fungi strains and non-infected leaves. Sensitivity experiments show that the primer PM1 an PM2 can detect target DNA at concentration of 0. 1 pg as well as the pathogen from the infected leaves 5 days after inoculation, The primers can be used for rapid and ac- curate detection of soybean downy mildew. The molecular detection method will be benefit of latent downy mildew infection and disease control measures carried out at early stage.

关 键 词：大豆霜霉病菌 ITS 分子探针 

分 类 号：S435.651[农业科学—农业昆虫与害虫防治]