辣椒疫霉菌RT-PCR检测技术的建立及应用

根据辣椒疫霉菌（Phytophthora capsici Leonian）与致病疫霉菌（Phytophthora infestans）及其他8种常见土传病害病原菌Actin基因序列差异，设计并筛选出辣椒疫霉菌的特异性引物YM2F/YM2R，建立辣椒疫霉菌的实时荧光定量PCR检测体系，并利用该体系定量检测人工模拟带菌样品及田间发病土壤样品中的辣椒疫霉菌。试验结果表明，利用该引物建立的实时荧光定量PCR检测体系线性关系良好，灵敏度为1 × 10-1 pg ? μL-1，是普通PCR的100倍。该体系无需病原菌的分离培养即可对土壤中的辣椒疫霉菌进行快速、特异且定量检测。对田间土壤样品的检测结果表明，在一定范围内病害发生、流行程度与病原菌密度成正比，从而为该病的流行监测和早期防控提供科学依据。

Development and Application of Real-time Fluorescent Quantitative PCR for Detection of Phytophthora capsici

According to the different bases of Actin gene sequences between Phytophthora capsici Leonian，P. infestans and other eight common soil pathogenic pathogens，we designed and screened out a specific primer pair YM2F/YM2R to establish a real-time fluorescent quantitative PCR detection assay of P. capsici，and using this assay to monitor P. capsici in soil samples prepared by artificial simulation and collecting form attacked yields. The results showed that the assay had a good liner relationship and a high sensitivity of 1 × 10-1 pg ? μL-1 DNA，which is 100 times more than normal PCR. P. capsici could be detected rapidly，specifically and quantitatively in infected soil without isolation or cultivation by the RT-PCR assay. The detection result of soil samples collected form attacked field indicated that the incidence and epidemiology of disease were proportional to the density of pathogenic bacteria within a certain range，which can provide scientific basis for the epidemic and early prevention of the disease.