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- Title
Effects of sgRNAs, Promoters, and Explants on the Gene Editing Efficiency of the CRISPR/Cas9 System in Chinese Kale.
- Authors
Huang, Wenli; Zheng, Aihong; Huang, Huanhuan; Chen, Zhifeng; Ma, Jie; Li, Xiangxiang; Liang, Qiannan; Li, Ling; Liu, Ruobin; Huang, Zhi; Qin, Yaoguo; Tang, Yi; Li, Huanxiu; Zhang, Fen; Wang, Qiaomei; Sun, Bo
- Abstract
The CRISPR/Cas9 system is extensively used for plant gene editing. This study developed an efficient CRISPR/Cas9 system for Chinese kale using multiple sgRNAs and two promoters to create various CRISPR/Cas9 vectors. These vectors targeted BoaZDS and BoaCRTISO in Chinese kale protoplasts and cotyledons. Transient transformation of Chinese kale protoplasts was assessed for editing efficiency at three BoaZDS sites. Notably, sgRNA: Z2 achieved the highest efficiency (90%). Efficiency reached 100% when two sgRNAs targeted BoaZDS with a deletion of a large fragment (576 bp) between them. However, simultaneous targeting of BoaZDS and BoaCRTISO yielded lower efficiency. Transformation of cotyledons led to Chinese kale mutants with albino phenotypes for boazds mutants and orange-mottled phenotypes for boacrtiso mutants. The mutation efficiency of 35S-CRISPR/Cas9 (92.59%) exceeded YAO-CRISPR/Cas9 (70.97%) in protoplasts, and YAO-CRISPR/Cas9 (96.49%) surpassed 35S-CRISPR/Cas9 (58%) in cotyledons. These findings introduce a strategy for enhancing CRISPR/Cas9 editing efficiency in Chinese kale.
- Subjects
CRISPRS; KALE; PLANT genes; COTYLEDONS; PROTOPLASTS; GENOME editing; GENETIC transformation; PHENOTYPES
- Publication
International Journal of Molecular Sciences, 2023, Vol 24, Issue 17, p13241
- ISSN
1661-6596
- Publication type
Article
- DOI
10.3390/ijms241713241