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Efficient DNA-free genome editing of bread wheat using CRISPR/Cas9 ribonucleoprotein complexes

Author

Listed:
  • Zhen Liang

    (State Key Laboratory of Plant Cell and Chromosome Engineering, Institute of Genetics and Developmental Biology, Chinese Academy of Sciences
    University of Chinese Academy of Sciences)

  • Kunling Chen

    (State Key Laboratory of Plant Cell and Chromosome Engineering, Institute of Genetics and Developmental Biology, Chinese Academy of Sciences)

  • Tingdong Li

    (State Key Laboratory of Plant Cell and Chromosome Engineering, Institute of Genetics and Developmental Biology, Chinese Academy of Sciences
    University of Chinese Academy of Sciences)

  • Yi Zhang

    (State Key Laboratory of Plant Cell and Chromosome Engineering, Institute of Genetics and Developmental Biology, Chinese Academy of Sciences
    University of Chinese Academy of Sciences)

  • Yanpeng Wang

    (State Key Laboratory of Plant Cell and Chromosome Engineering, Institute of Genetics and Developmental Biology, Chinese Academy of Sciences
    University of Chinese Academy of Sciences)

  • Qian Zhao

    (State Key Laboratory of Plant Cell and Chromosome Engineering, Institute of Genetics and Developmental Biology, Chinese Academy of Sciences
    University of Chinese Academy of Sciences)

  • Jinxing Liu

    (State Key Laboratory of Plant Cell and Chromosome Engineering, Institute of Genetics and Developmental Biology, Chinese Academy of Sciences)

  • Huawei Zhang

    (State Key Laboratory of Plant Cell and Chromosome Engineering, Institute of Genetics and Developmental Biology, Chinese Academy of Sciences)

  • Cuimin Liu

    (State Key Laboratory of Plant Cell and Chromosome Engineering, Institute of Genetics and Developmental Biology, Chinese Academy of Sciences)

  • Yidong Ran

    (Genovo Biotechnology Co., Ltd)

  • Caixia Gao

    (State Key Laboratory of Plant Cell and Chromosome Engineering, Institute of Genetics and Developmental Biology, Chinese Academy of Sciences)

Abstract

Substantial efforts are being made to optimize the CRISPR/Cas9 system for precision crop breeding. The avoidance of transgene integration and reduction of off-target mutations are the most important targets for optimization. Here, we describe an efficient genome editing method for bread wheat using CRISPR/Cas9 ribonucleoproteins (RNPs). Starting from RNP preparation, the whole protocol takes only seven to nine weeks, with four to five independent mutants produced from 100 immature wheat embryos. Deep sequencing reveals that the chance of off-target mutations in wheat cells is much lower in RNP mediated genome editing than in editing with CRISPR/Cas9 DNA. Consistent with this finding, no off-target mutations are detected in the mutant plants. Because no foreign DNA is used in CRISPR/Cas9 RNP mediated genome editing, the mutants obtained are completely transgene free. This method may be widely applicable for producing genome edited crop plants and has a good prospect of being commercialized.

Suggested Citation

  • Zhen Liang & Kunling Chen & Tingdong Li & Yi Zhang & Yanpeng Wang & Qian Zhao & Jinxing Liu & Huawei Zhang & Cuimin Liu & Yidong Ran & Caixia Gao, 2017. "Efficient DNA-free genome editing of bread wheat using CRISPR/Cas9 ribonucleoprotein complexes," Nature Communications, Nature, vol. 8(1), pages 1-5, April.
    • Handle: RePEc:nat:natcom:v:8:y:2017:i:1:d:10.1038_ncomms14261
    DOI: 10.1038/ncomms14261
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    Citations

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    Cited by:

    1. Hang Su & Yuanchun Wang & Jin Xu & Ahmad A. Omar & Jude W. Grosser & Milica Calovic & Liyang Zhang & Yu Feng & Christopher A. Vakulskas & Nian Wang, 2023. "Generation of the transgene-free canker-resistant Citrus sinensis using Cas12a/crRNA ribonucleoprotein in the T0 generation," Nature Communications, Nature, vol. 14(1), pages 1-12, December.
    2. You-Jeong Kim & Dayoung Yun & Jungjoon K. Lee & Cheulhee Jung & Aram J. Chung, 2024. "Highly efficient CRISPR-mediated genome editing through microfluidic droplet cell mechanoporation," Nature Communications, Nature, vol. 15(1), pages 1-12, December.

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