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Cas13a-based multiplex RNA targeting for potato virus Y

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Abstract

Main conclusion

The Cas13a-based multiplex RNA targeting system can be engineered to confer resistance to RNA viruses, whereas the number and expression levels of gRNAs have no significant effect on viral interference.

Abstract

The CRISPR–Cas systems provide adaptive immunity to bacterial and archaeal species against invading phages and foreign plasmids. The class 2 type VI CRISPR/Cas effector Cas13a has been harnessed to confer the protection against RNA viruses in diverse eukaryotic species. However, whether the number and expression levels of guide RNAs (gRNAs) have effects on the efficiency of RNA virus inhibition is unknown. Here, we repurpose CRISPR/Cas13a in combination with an endogenous tRNA-processing system (polycistronic tRNA–gRNA) to target four genes of potato virus Y (PVY) with varying expression levels. We expressed Cas13a and four different gRNAs in potato lines, and the transgenic plants expressing multiple gRNAs displayed similar suppression of PVY accumulation and reduced disease symptoms as those expressing a single gRNA. Moreover, PTG/Cas13a-transformed plants with different expression levels of multiple gRNAs displayed similar resistance to PVY strains. Collectively, this study suggests that the Cas13a-based multiplex RNA targeting system can be utilized to engineer resistance to RNA viruses in plants, whereas the number and expression levels of gRNAs have no significant effect on CRISPR/Cas13a-mediated viral interference in plants.

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Data availability

Data sharing not applicable to this article as no datasets were generated or analyzed during the current study. All data generated or analyzed during this study are included in this article (and its supplementary information files).

Abbreviations

PVY:

Potato virus Y

gRNA:

Guide RNA

ELISA:

Enzyme-linked immunosorbent assay

dpi:

Days post-infection

PTG:

Polycistronic tRNA–gRNA

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Acknowledgements

This work was supported by Grants from the National Natural Science Foundation of China (32271912 and 32271546) and the Natural Science Foundation of Hubei Province (2022CFB482).

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Author notes

  1. Xiaohui Zhan and Zhen Tu contributed equally to this work.

Authors and Affiliations

  1. State Key Laboratory of Biocatalysis and Enzyme Engineering, School of Life Sciences, Hubei Hongshan Laboratory, Hubei University, Wuhan, 430062, China

    Xiaohui Zhan, Wenlei Song, Shengchun Li, Jiang Zhang & Fengjuan Zhang

  2. Key Laboratory of Potato Biology and Biotechnology, Key Laboratory of Horticultural Plant Biology, Huazhong Agricultural University, Wuhan, 430070, China

    Zhen Tu & Bihua Nie

  3. Agricultural Genomics Institute at Shenzhen, Chinese Academy of Agricultural Sciences, Shenzhen, China

    Jiang Zhang

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  1. Xiaohui Zhan
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Correspondence to Fengjuan Zhang.

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Zhan, X., Tu, Z., Song, W. et al. Cas13a-based multiplex RNA targeting for potato virus Y. Planta 258, 70 (2023). https://doi.org/10.1007/s00425-023-04216-x

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