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KMID : 1098420230310060359
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Korean Journal of Medicinal Crop Science
2023 Volume.31 No. 6 p.359 ~ p.370
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Development of Lateral Flow Reverse Transcription Recombinase Polymerase Amplification (LF-RT-RPA) Assay for On-site Detection of Cucumber Mosaic Virus on Cnidium officinale
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Lee Hyun-Taek
Yoon Ju-Yeon
Ju Ho-Jong
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Abstract
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Background: Quality and quantity of Cheongung (Cnidium officinale Makinoi) is reduced by plant viral infections. Therefore, it is necessary to develop effective detection methods for diagno- sis of problematic viruses in Cheongung farms. This study aimed to develop lateral flow reverse transcription recombinase polymerase amplification (LF-RT-RPA) assay for the detection of cucumber mosaic virus (CMV) in C. officinale.
Methods and Results: Total RNA was extracted from CMV-infected Cheongung plant and sub- jected to RT-RPA reaction with RPA_CMV-CP-F1/R1 primers targeting the CMV RNA3 sequence. When specificity was examined using lateral flow Immunostrip, experimental and con- trol lines were generated only in the sample from the CMV-infected plant, while control lines were observed in samples from cnidium vein yellowing virus-1, cnidium vein yellowing virus-2, and apple stem grooving virus-infected Cheongung plants. The optimal temperature and reaction time for LF-RT-RPA assay were determined to be 37oC and 15 min. On-site diagnosis carried out to detect CMV in 7 Cheongung plants showing viral-infection symptoms, confirmed six of the plants infected with CMV.
Conclusion: CMV, one of the prevalent plant viruses, was effectively detected using LF-RT-RPA assay without the need of any equipment on farms. Detection limit for CMV in C. officinale Maki- noi was up to 100 fg of total RNA, suggesting that the LF-RT-RPA assay developed in this study is suitable for on-site detection of CMV in Cheongung farms.
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KEYWORD
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Cnidium officinale Makino, Cucumber mosaic virus, Detection, Lateral Flow Immunoassay, On-site, Reverse Transcription-Recombinase Polymerase Amplification
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