The Plant Pathology Journal

한국식물병리학회 (The Korean Society of Plant Pathology)
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Development and Application of Reverse Transcription Nanoplate-Based Digital PCR Assay for Sensitive and Accurate Detection of Rice Black-Streaked Dwarf Virus in Cereal Crops

  • Hyo-Jeong Lee (Department of Applied Biology, Chonnam National University) ;
  • Hae-Jun Kim (Department of Applied Biology, Chonnam National University) ;
  • Sang-Min Kim (Crop Foundation Research Division, National Institute of Crop Science, Rural Development Administration) ;
  • Rae-Dong Jeong (Department of Applied Biology, Chonnam National University)
  • 투고 : 2024.03.05
  • 심사 : 2024.06.25
  • 발행 : 2024.08.01
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초록

The emergence of rice black-streaked dwarf virus (RBSDV) poses a significant threat to global cereal crop cultivation, necessitating the urgent development of reliable detection and quantification techniques. This study introduces a reliable approach for the precise and sensitive quantification of the RBSDV in cereal crop samples, employing a reverse transcription digital polymerase chain reaction (RT-dPCR) assay. We assessed the specificity and sensitivity of the RT-dPCR assay proposed for precise RBSDV detection and quantification. Our findings demonstrate that RT-dPCR was specific for detection of RBSDV, with no cross-reactivity observed with other viruses infecting cereal crops. The RT-dPCR sensitivity was over 10 times that of RT-quantitative PCR (RT-qPCR). The detection limit of RT-dPCR was 0.096 copies/㎕. In addition, evaluation of RT-dPCR assay with field samples was conducted on 60 different cereal crop samples revealed that RT-dPCR (45/60) exhibited superior accuracy compared with RT-qPCR (23/60). In this study, we present a specific and accurate RT-dPCR assay for the detection and quantification of RBSDV.

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과제정보

This work was carried out with the support of Cooperative Research Program for Agriculture Science and Technology Development (Project No. PJ014995022023) Rural Development Administration, Republic of Korea.

참고문헌

  1. Hua, Y., Feng, C., Gu, T., Chen, H., Liu, D., Xu, K. and Zhang, K. 2023. Development of polyclonal antibodies and a serological-based reverse-transcription loop-medaited isothermal amplification (S-RT-LAMP) assay for rice balck-streaked dwarf virus detection in both rice and small brown planthopper. Viruses 15:2127.
  2. Kim, J.-S., Lee, S.-H., Choi, H.-S., Cho, J.-D., Noh, T.-W. and Kim, J.-Y. 2009. Convenient genetic diagnosis of virion captured (VC)/RT-PCR for rice viruses (RSV, RBSDV) and small brown plant hopper. Res. Plant Dis. 15:57-62 (in Korean). https://doi.org/10.5423/RPD.2009.15.2.057
  3. Kim, N.-K., Lee, H.-Y., Kim, S.-M. and Jeong, R.-D. 2022a. Identification of viruses infecting oats in Korea by metatranscriptomics. Plants 11:256.
  4. Kim, S.-W., Lee, H.-J., Cho, K.-H. and Jeong, R.-D. 2022b. Detection and quantification of apple stem grooving virus in micropropagated apple plantlets using reverse-transcription droplet digital PCR. Plant Pathol. J. 38:417-422. https://doi.org/10.5423/PPJ.NT.05.2022.0072
  5. Kim, S.-W., Lee, H.-J., Cho, S.-Y. and Jeong, R.-D. 2023. Development and preliminary application of a reverse-transcription droplet digital PCR assay for detection and quantification of apple pitting virus in vitro propagated apple plantlets. Sci. Hortic. 321:112363.
  6. Lee, B.-C., Hong, Y.-K., Hong, S.-J., Park, S.-T. and Lee, K.-W. 2005. Occurrence and detection of rice black-streaked dwarf virus in Korea. Plant Pathol. J. 21:172-173. https://doi.org/10.5423/PPJ.2005.21.2.172
  7. Lee, H.-J., Cho, I.-S., Ju, H.-J. and Jeong, R.-D. 2021. Development of a reverse transcription droplet digital PCR assay for sensitive detection of peach latent mosaic viroid. Mol. Cell. Probes 58:101746.
  8. Lei, S., Chen, S. and Zhong, Q. 2021. Digital PCR for accurate quantificaiton of pathogens: principles, applications, challenges and future prospects. Int. J. Biol. Macromol. 184:750-759. https://doi.org/10.1016/j.ijbiomac.2021.06.132
  9. Wang, P., Liu, J., Lyu, Y., Huang, Z., Zhang, X., Sun, B., Li, P., Jing, X., Li, H. and Zhang, C. 2022. A review of vector-borne rice viruse. Viruses 14:2258.
  10. Wu, N., Zhang, L., Ren, Y. and Wang, X. 2020. Rice black-streaked dwarf virus: from multiparty interactions among plant-virus-vector to intermittent epidemics. Mol. Plant Pathol. 21:1007-1019. https://doi.org/10.1111/mpp.12946
  11. Zhang, P., Mar, T. T., Liu, W., Li, L. and Wang, X. 2013. Simultaneous dection and differentiation of rice black streaked dwarf virus (RBSDV) and southern rice balck streaked dwarf virus (SRBSDV) by duplex real time RT-PCR. Virol. J. 10:24.
  12. Zhao, C., Sun, F., Li, X., Lan, Y., Du, L., Zhou, T. and Zhou, Y. 2019. Reverse transcription-recombinase polymerase amplification combined with lateral flow strip for detection of rice black-streaked dwarf virus in plants. J. Virol. Methods 263:96-100. https://doi.org/10.1016/j.jviromet.2018.11.001