PubMed 34782773

PubMed ID: 34782773

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A donor-DNA-free CRISPR/Cas-based approach to gene knock-up in rice.
Authors: Lu Yu, Wang Jiyao, Chen Bo, Mo Sudong, Lian Lei, Luo Yanmin, Ding Dehui, Ding Yanhua, Cao Qing, Li Yucai, Li Yong, Liu Guizhi, Hou Qiqi, Cheng Tingting, Wei Junting, Zhang Yanrong, Chen Guangwu, Song Chao, Hu Qiang, Sun Shuai, Fan Guangyi, Wang Yating, Liu Zhiting, Song Baoan, Zhu Jian-Kang, Li Huarong, Jiang Linjian
Journal: Nature plants (Nat Plants), Vol.7(11), 2021‑Nov

DOI: 10.1093/database/baaa055 PMCID: 5827348

Abstract
Structural variations (SVs), such as inversion and duplication, contribute to important agronomic traits in crops1. Pan-genome studies revealed that SVs were a crucial and ubiquitous force driving genetic diversification2-4. Although genome editing can effectively create SVs in plants and animals5-8, the potential of designed SVs in breeding has been overlooked. Here, we show that new genes and traits can be created in rice by designed large-scale genomic inversion or duplication using CRISPR/Cas9. A 911 kb inversion on chromosome 1 resulted in a designed promoter swap between CP12 and PPO1, and a 338 kb duplication between HPPD and Ubiquitin2 on chromosome 2 created a novel gene cassette at the joint, promoterUbiquitin2::HPPD. Since the original CP12 and Ubiquitin2 genes were highly expressed in leaves, the expression of PPO1 and HPPD in edited plants with homozygous SV alleles was increased by tens of folds and conferred sufficient herbicide resistance in field trials without adverse effects on other important agronomic traits. CRISPR/Cas-based genome editing for gene knock-ups has been generally considered very difficult without inserting donor DNA as regulatory elements. Our study challenges this notion by providing a donor-DNA-free strategy, thus greatly expanding the utility of CRISPR/Cas in plant and animal improvements.
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