Rice false smut caused by Ustilaginoidea virens is one of the most devastating diseases on rice worldwide, which results in serious reduction of rice quality and yield. As an airborne fungal disease, early diagnosis of rice false smut, monitoring the epidemics and distribution of its pathogens, is particularly important to management the infection. In this study, a quantitative loop-mediated isothermal amplification (q-LAMP) method for the U. virens detection and quantifying was developed. This method has higher sensitivity and efficiency compared to quantitative real-time PCR (q-PCR) method. The species-specific primer sets UV-2 used was design based on the unique sequence of U. virens ustiloxins biosynthetic gene (NCBI accession number: BR001221.1). The q-LAMP assay was able to detect a concentration of 6.4 spores/mL at an optimal reaction temperature of 63.4℃ within 60 min. Moreover, the q-LAMP assay can even achieve accurate quantitative detection when there were only 9 spores on the tape. A linearized equation for the standard curve, y =-0.2866x + 13.829 (x is the amplification time, the spore number = 100.65y), was established for detection and quantifying of U. virens. In field detection applications, this q-LAMP method is more accurate and faster than traditional observation method. Collectively, this study has established a powerful and simple monitoring tool for U. virens, which provide valuable technical supports for forecast and management of rice false smut, and theoretical basis for precise fungicide application.
Biology and Life Sciences, Biochemistry and Molecular Biology
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