文献类型： 外文期刊

作者： Li, Yanli ¹ ; Huang, Changjun ² ; Liu, Yong ² ; Zeng, Jianmin ² ; Yu, Haiqin ² ; Tong, Zhijun ² ; Yuan, Xinjie ³ ; Sui, Xueyi ² ; Fang, Dunhuang ² ; Xiao, Bingguang ² ; Zhao, Shancen ¹ ; Yuan, Cheng ² ;

作者机构： 1.BGI Shenzhen, Shenzhen, Guangdong, Peoples R China

2.Yunnan Acad Tobacco Agr Sci, Natl Tobacco Genet Engn Res Ctr, Kunming, Yunnan, Peoples R China

3.Jiangxi Acad Agr Sci, Inst Vegetables & Flowers, Nanchang, Peoples R China

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.