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Effects of sgRNAs, Promoters, and Explants on the Gene Editing Efficiency of the CRISPR/Cas9 System in Chinese Kale
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 cotyledo...
Autores principales: | , , , , , , , , , , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
MDPI
2023
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10487834/ https://www.ncbi.nlm.nih.gov/pubmed/37686051 http://dx.doi.org/10.3390/ijms241713241 |
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author | 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 |
author_facet | 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 |
author_sort | Huang, Wenli |
collection | PubMed |
description | 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. |
format | Online Article Text |
id | pubmed-10487834 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2023 |
publisher | MDPI |
record_format | MEDLINE/PubMed |
spelling | pubmed-104878342023-09-09 Effects of sgRNAs, Promoters, and Explants on the Gene Editing Efficiency of the CRISPR/Cas9 System in Chinese Kale 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 Int J Mol Sci Article 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. MDPI 2023-08-26 /pmc/articles/PMC10487834/ /pubmed/37686051 http://dx.doi.org/10.3390/ijms241713241 Text en © 2023 by the authors. https://creativecommons.org/licenses/by/4.0/Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/). |
spellingShingle | Article 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 Effects of sgRNAs, Promoters, and Explants on the Gene Editing Efficiency of the CRISPR/Cas9 System in Chinese Kale |
title | Effects of sgRNAs, Promoters, and Explants on the Gene Editing Efficiency of the CRISPR/Cas9 System in Chinese Kale |
title_full | Effects of sgRNAs, Promoters, and Explants on the Gene Editing Efficiency of the CRISPR/Cas9 System in Chinese Kale |
title_fullStr | Effects of sgRNAs, Promoters, and Explants on the Gene Editing Efficiency of the CRISPR/Cas9 System in Chinese Kale |
title_full_unstemmed | Effects of sgRNAs, Promoters, and Explants on the Gene Editing Efficiency of the CRISPR/Cas9 System in Chinese Kale |
title_short | Effects of sgRNAs, Promoters, and Explants on the Gene Editing Efficiency of the CRISPR/Cas9 System in Chinese Kale |
title_sort | effects of sgrnas, promoters, and explants on the gene editing efficiency of the crispr/cas9 system in chinese kale |
topic | Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10487834/ https://www.ncbi.nlm.nih.gov/pubmed/37686051 http://dx.doi.org/10.3390/ijms241713241 |
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