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DNA-free genome editing in plants with preassembled CRISPR-Cas9 ribonucleoproteins

Nature Biotechnology volume 33, pages 11621164 (2015) | Download Citation

Abstract

Editing plant genomes without introducing foreign DNA into cells may alleviate regulatory concerns related to genetically modified plants. We transfected preassembled complexes of purified Cas9 protein and guide RNA into plant protoplasts of Arabidopsis thaliana, tobacco, lettuce and rice and achieved targeted mutagenesis in regenerated plants at frequencies of up to 46%. The targeted sites contained germline-transmissible small insertions or deletions that are indistinguishable from naturally occurring genetic variation.

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Acknowledgements

This work was supported in part by grants from the Institute for Basic Science (IBS-R021-D1) and the Next-Generation BioGreen21 Program (PJ01104501 to S.C. and PJ01104502 to S.I.K.).

Author information

Author notes

    • Seung Woo Cho

    Present address: Program in Epithelial Biology, Stanford University School of Medicine, Stanford, California, USA.

    • Je Wook Woo
    •  & Jungeun Kim

    These authors contributed equally to this work.

Affiliations

  1. Convergence Research Center for Functional Plant Products, Advanced Institutes of Convergence Technology, Yeongtong-gu, Suwon-si, Gyeonggi-do, Korea.

    • Je Wook Woo
    • , Soon Il Kwon
    •  & Sunghwa Choe
  2. Center for Genome Engineering, Institute for Basic Science, Seoul, South Korea.

    • Jungeun Kim
    • , Hyeran Kim
    • , Sang-Gyu Kim
    • , Sang-Tae Kim
    •  & Jin-Soo Kim
  3. Department of Chemistry, Seoul National University, Seoul, South Korea.

    • Jungeun Kim
    • , Seung Woo Cho
    •  & Jin-Soo Kim
  4. School of Biological Sciences, College of Natural Sciences, Seoul National University, Seoul, South Korea.

    • Claudia Corvalán
    •  & Sunghwa Choe
  5. Plant Genomics and Breeding Institute, Seoul National University, Seoul, South Korea.

    • Sunghwa Choe

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Contributions

J.-S.K. and S.C. supervised the research. J.W.W., S.I.K. and C.C. carried out plant regeneration. J.K., S.W.C. H.K., S.-G.K. and S.-T.K. performed mutation analysis.

Competing interests

J.-S.K. and S.C. are co-inventors on a patent application covering the genome editing method described in this manuscript.

Corresponding authors

Correspondence to Sunghwa Choe or Jin-Soo Kim.

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DOI

https://doi.org/10.1038/nbt.3389

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