CRISPR/Cas9-mediated seamless gene replacement in protoplasts expands the resistance spectrum to TMV-U1 strain in regenerated Nicotiana tabacum

文献类型： 外文期刊

第一作者： Li, Yanli

作者： Li, Yanli;Zhao, Shancen;Huang, Changjun;Liu, Yong;Zeng, Jianmin;Yu, Haiqin;Tong, Zhijun;Sui, Xueyi;Fang, Dunhuang;Xiao, Bingguang;Yuan, Cheng;Yuan, Xinjie

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关键词： protoplast regeneration; gene knock-in; gene replacement; CRISPR/Cas9; long fragment insertion; TMV resistance

期刊名称：PLANT BIOTECHNOLOGY JOURNAL （ 影响因子：13.8； 五年影响因子：13.2 ）

ISSN： 1467-7644

年卷期： 2023 年

页码：

收录情况： SCI

摘要： CRISPR/Cas-based genome editing is now extensively used in plant breeding and continues to evolve. Most CRISPR/Cas current applications in plants focus on gene knock-outs; however, there is a pressing need for new methods to achieve more efficient delivery of CRISPR components and gene knock-ins to improve agronomic traits of crop cultivars. We report here a genome editing system that combines the advantages of protoplast technologies with recent CRISPR/Cas advances to achieve seamless large fragment insertions in the model Solanaceae plant Nicotiana tabacum. With this system, two resistance-related regions of the N' gene were replaced with homologous fragments from the N' alata gene to confer TMV-U1 resistance in the T0 generation of GMO-free plants. Our study establishes a reliable genome-editing tool for efficient gene modifications and provides a detailed description of the optimization process to assist other researchers adapt this system for their needs.

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