Establishment and Application of Real-time Quantitative PCR Detection System for the Pathogenic Fusarium oxysporum Causing Astragalus membranaceus var. mongholicus Root Rot
1 Modern Research Center for Traditional Chinese Medicine, Shanxi University, Taiyuan 030006, China; 2 Institute of the Industrial Crop, Shanxi Agricultural University, Taiyuan 030031, China; 3 Institute of Applied Chemistry, Shanxi University, Taiyuan 030006, China
Abstract：Fusarium oxysporum is one of the dominant pathogens causing Astragalus membranaceus var. mongholicus (AMM) root rot. In order to rapidly and accurately detect and quantify F. oxysporum in AMM plants and soil, primers were designed based on the elongation factor-1α (EF-1α) sequence and verified for their specificity. The real-time quantitative PCR (qRT-PCR) detection system was established for the pathogen. The constructed qRT-PCR system was used to detect F. oxysporum in AMM plants and soil from AMM planting sites. The results showed that primer pair FaeF2/FaeR2 was highly specific, and the sensitivities of qRT-PCR detection were 0.896 pg/μL for F. oxysporum genomic DNA in AMM and 102 conidia/g for F. oxysporum conidia in soil. The correlation coefficients of the constructed standard curves for F. oxysporum detection in AMM and soil were 0.998 and 0.973, respectively. Repeatability evaluation showed that the system was stable and reliable. The AMM plants and soil samples were detected by the qRT-PCR detection system. The detection rate of the pathogenic F. oxysporum was 100% in F. oxysporum-inoculated samples, 66.7% in suspected diseased samples and 43.75% in soil samples; the pathogen colonization in AMM was positively related with the disease index and the number of conidia in diseased soil was significantly greater than that in healthy soil at the same AMM cultivation years. The qRT-PCR system established in this study was suitable for the detection of the pathogenic F. oxysporum causing AMM root rot, and will provide technical support for the rapid diagnosis, accurate evaluation and soil warning of AMM root rot.
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