Abstract
Fig mosaic virus (FMV), a negative-strand RNA virus, is a causal agent of fig mosaic disease, which occurs in almost all countries where figs are grown and severely affects worldwide fig production. Reverse transcription-polymerase chain reaction (RT-PCR) using FMV-specific primers has typically been used to detect FMV, but faster and easier detection methods for FMV are required because RT-PCR requires multiple steps and special equipment. In this study, we established a reverse transcription loop-mediated isothermal amplification (RT-LAMP) method to detect FMV. We designed LAMP primer sets based on sequence conservation among FMV isolates. The RT-LAMP reaction using a primer set targeting RNA3 in total RNA extracted from FMV-infected fig leaves resulted in rapid detection within 15 min. In addition, we established a fast and simple method of direct sampling from fig leaves using a wooden toothpick. The RT-LAMP reaction specificity and reactivity when using the direct sampling method were almost comparable to those obtained using isolated total RNA. Moreover, geographically and phylogenetically distinct FMV isolates were detectable using the FMV RT-LAMP assay. The assay presented here provides a practical method to detect FMV.
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Acknowledgments
This work was supported by Grants-in-Aid for Scientific Research from the Japan Society for the Promotion of Science. We thank Bojan Duduk (Institute of Pesticides and Environmental Protection, Belgrade-Zemun, Serbia), Shimane Agricultural Technology Center (Shimane, Japan) and Fukuoka Prefectural Plant Protection Office (Fukuoka, Japan) for kindly providing FMV-infected fig samples.
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Ishikawa, K., Maejima, K., Netsu, O. et al. Rapid detection of fig mosaic virus using reverse transcription loop-mediated isothermal amplification. J Gen Plant Pathol 81, 382–389 (2015). https://doi.org/10.1007/s10327-015-0603-1
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DOI: https://doi.org/10.1007/s10327-015-0603-1