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Virus-induced CRISPR-Cas9 system improved resistance against tomato yellow leaf curl virus

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Abstract

Plant viruses are the most significant factors associated with massive economical losses in agricultural industries worldwide. Accordingly, many studies are dedicated to making virus-resistant crop varieties each year due to the ever-changing nature of viruses. Recently genome engineering methods have been used to confer interference against eukaryotic viruses. Research results on genome editing technics, in particular, CRISPR-Cas9, promises a feasible solution to make virus-resistant crops. In this research, we explored the possibility of utilizing CRISPR-Cas9 to obtain TYLCV resistant tomato varieties. Moreover, to overcome any potential off-target effects of Cas9, we used an inducible promoter to initiate Cas9 activity in case of the virus attack. Cas9 vector was transformed by the rgsCaM promoter, known as an endogenous silencer of RNAi and overexpressed after a virus attack. The golden gate cloning method was applied to construct sgRNAs. Intergenic region and coat protein-coding sequences of TYLCV were used to design sgRNAs. Infiltrated sensitive Money Maker varieties analyzed by real-time PCR, showed a significant reduction or delayed accumulation of viral DNA compared to the control plants. This result demonstrates the efficiency of using an inducible promoter in CRISPR-Cas9 constructs.

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Acknowledgements

This work was financially supported by grant Number 950709 of the Biotechnology Development Council of the Islamic Republic of Iran.

Funding

This study was funded by Ferdowsi University of Mashhad (Grant No. 3/41501) and the Biotechnology Council of the Islamic Republic of Iran (Grant No. 950709).

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Authors and Affiliations

  1. Department of Biotechnology and Plant Breeding, Faculty of Agriculture, Ferdowsi University of Mashhad, Mashhad, Iran

    Parisa Ghorbani Faal, Mohammad Farsi, Alireza Seifi & Amin Mirshamsi Kakhki

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  1. Parisa Ghorbani Faal
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  2. Mohammad Farsi
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  3. Alireza Seifi
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  4. Amin Mirshamsi Kakhki
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Correspondence to Mohammad Farsi.

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Conflict of interest

Author Mohammad Farsi has received research grants from Ferdowsi University. Author Parisa Ghorbani Faal has received research grants from the Biotechnology Development Council of the Islamic Republic of Iran. Author Alireza Seifi declares that he has no conflict of interest. Author Amin Mirshamsi Kakhki declares that he has no conflict of interest.

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Ghorbani Faal, P., Farsi, M., Seifi, A. et al. Virus-induced CRISPR-Cas9 system improved resistance against tomato yellow leaf curl virus. Mol Biol Rep 47, 3369–3376 (2020). https://doi.org/10.1007/s11033-020-05409-3

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  • DOI: https://doi.org/10.1007/s11033-020-05409-3

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