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- Title
Generation of the transgene-free canker-resistant Citrus sinensis using Cas12a/crRNA ribonucleoprotein in the T0 generation.
- Authors
Su, Hang; Wang, Yuanchun; Xu, Jin; Omar, Ahmad A.; Grosser, Jude W.; Calovic, Milica; Zhang, Liyang; Feng, Yu; Vakulskas, Christopher A.; Wang, Nian
- Abstract
Citrus canker caused by Xanthomonas citri subsp. citri (Xcc) is a destructive citrus disease worldwide. Generating disease-resistant cultivars is the most effective, environmentally friendly and economic approach for disease control. However, citrus traditional breeding is lengthy and laborious. Here, we develop transgene-free canker-resistant Citrus sinensis lines in the T0 generation within 10 months through transformation of embryogenic protoplasts with Cas12a/crRNA ribonucleoprotein to edit the canker susceptibility gene CsLOB1. Among the 39 regenerated lines, 38 are biallelic/homozygous mutants, demonstrating a 97.4% biallelic/homozygous mutation rate. No off-target mutations are detected in the edited lines. Canker resistance of the cslob1-edited lines results from both abolishing canker symptoms and inhibiting Xcc growth. The transgene-free canker-resistant C. sinensis lines have received regulatory approval by USDA APHIS and are exempted from EPA regulation. This study provides a sustainable and efficient citrus canker control solution and presents an efficient transgene-free genome-editing strategy for citrus and other crops. Development of canker-resistant citrus cultivars via traditional approaches is a lengthy and laborious process. Here, the authors report the generation of regulatory approval, transgene-free, canker-resistant sweet orange lines using Cas12a/crRNA ribonucleoprotein-based susceptibility gene editing strategy.
- Subjects
UNITED States. Dept. of Agriculture; UNITED States. Environmental Protection Agency; CANKER (Plant disease); CITRUS canker; XANTHOMONAS campestris; CITRUS; ORANGES; REGULATORY approval; GENOME editing; PROTOPLASTS
- Publication
Nature Communications, 2023, Vol 14, Issue 1, p1
- ISSN
2041-1723
- Publication type
Article
- DOI
10.1038/s41467-023-39714-9