烟草青枯病、黑胫病和猝倒病的多重PCR检测

Triplex PCR Detection of Ralstonia solanacearum,Phytophthora nicotianae and Pythium aphanidermatum from Infected Tobacco Tissues

烟草青枯病、黑胫病和猝倒病是烟草生产中的重要病害,常发生复合侵染。以3种重要烟草病原菌基因组DNA作为目的基因,分别设计3对特异性引物,通过优化引物和模板浓度,摸索扩增参数,建立了能同时检测3种病害的多重聚合酶链式反应(Multiplex polymerase chain reaction,mPCR)体系。结果表明,多重PCR能分别扩增出长度为461,364,265 bp的特异性目的条带,能同时检测3种病原菌DNA的最低检出限为1.01 ng/μL,该多重PCR检测体系反应准确、快速、高效,为同时检测以上3种烟草病害提供了新的方法,对烟草复合侵染病害的诊断和防治提供了重要的指导作用。

R. solanacearum,P. nicotianae and P. aphanidermatum are important compound infection diseases of the tobacco production. In this study,Three pairs of primers were designed respectively according to the genomic DNA of R. solanacearum,P. nicotiana and P. aphanidermatum. And they were used to amplify simultaneously the DNA sequences by multiplex PCR and the reaction conditions were optimized. The results showed that the multiplex PCR assay amplified three specific target channels of 461,364 and 265 bp. The detection limits of the multiplex PCR for R. solanacearum,P. nicotiana and P. aphanidermatum were 1. 01 ng / μL. This multiplex PCR method was accurate,fast and effective and provided a new approach and an important guiding role for the tobacco disease compound infection diagnosis and treatment.