Establishment and application of RT-LAMP method for rapid detection of Rehmannia mosaic virus
Based on reverse transcription loop-mediated isothermal amplification(RT-LAMP)technology,a rapid,specific and sensitive RT-LAMP detection method for Rehmannia mosaic virus(ReMV)was established.Three sets of LAMP primers were designed according to the nucleotide sequence of ReMV coat protein(CP),and the optimized primers were determined by primer screening.Temperature optimization,specificity detection and sensitivity comparison of RT-LAMP were carried out by using electrophoresis detection and visual(SYBR Green Ⅰchromogenic agent).The results showed that the optimum detection temperature of RT-LAMP method was 60℃.ReMV could be specifically detected by the optimized LAMP,and the sensitivity was 1 000 times higher than that of conventional PCR.It could be detected in 1.2 × 10-1 copies/pL.The detection results of 60 samples of Rehmannia glutinosa showed that the positive detection rate of RT-LAMP was 98.3％,which was higher than that of conventional RT-PCR(75％).The RT-LAMP detection technology established in this study provides a fast and efficient method for the accurate detection of ReMV.
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