J Integr Plant Biol ›› 2023, Vol. 65 ›› Issue (4): 900-906.DOI: 10.1111/jipb.13428

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1†   

  • 收稿日期:2022-10-06 接受日期:2022-12-07 出版日期:2023-04-01 发布日期:2023-04-08

Optimized prime editing efficiently generates heritable mutations in maize

Dexin Qiao1†, Junya Wang1†, Min‐Hui Lu2†, Cuiping Xin1, Yiping Chai1, Yuanyuan Jiang1, Wei Sun1, Zhenghong Cao1, Siyi Guo3, Xue‐Chen Wang1 and Qi‐Jun Chen1,2*   

  1. 1. State Key Laboratory of Plant Physiology and Biochemistry, College of Biological Sciences, China Agricultural University, Beijing 100193, China;
    2. Center for Crop Functional Genomics and Molecular Breeding, China Agricultural University, Beijing 100193, China;
    3. Collaborative Innovation Center of Crop Stress Biology, Henan Province, Institute of Plant Stress Biology, School of Life Science, Henan University, Kaifeng 475004, China
    †These authors contributed equally to this work.
    *Correspondence: Qi‐Jun Chen (qjchen@cau.edu.cn)
  • Received:2022-10-06 Accepted:2022-12-07 Online:2023-04-01 Published:2023-04-08

Abstract: Low efficiency is the main obstacle to using prime editing in maize (Zea mays). Recently, prime‐editing efficiency was greatly improved in mammalian cells and rice (Oryza sativa) plants by engineering prime‐ editing guide RNAs (pegRNAs), optimizing the prime editor (PE) protein, and manipulating cellular determinants of prime editing. In this study, we tested PEs optimized via these three strategies in maize. We demonstrated that the ePE5max system, composed of PEmax, epegRNAs (pegRNA‐ evopreQ. 1), nicking single guide RNAs (sgRNAs), and MLH1dn, efficiently generated heritable mutations that conferred resistance to herbicides that inhibit 5‐enolpyruvylshikimate‐3‐phosphate synthase (EPSPS), acetolactate synthase (ALS), or acetyl CoA carboxylase (ACCase) activity. Collectively, we demonstrate that the ePE5max system has sufficient efficiency to generate heritable (homozygous or heterozygous) mutations in maize target genes and that the main obstacle to using PEs in maize has thus been removed.

Key words: CRISPR/Cas, epegRNAs, herbicide resistance, maize, MLH1dn, prime editing

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