Editing the RR-TZF Gene Subfamily in Rice Uncovers Potential Risks of CRISPR/Cas9 for Targeted Genetic Modification

文献类型： 外文期刊

第一作者： Zhou, Shufen

作者： Zhou, Shufen;Lu, Xiaozhuan;Wu, Weiren;Zhou, Shufen;Tian, Dagang;Liu, Huaqing;Zhang, Di;Chen, Rui;Yang, Shaohua;Wang, Feng

作者机构：

关键词： CRISPR/Cas9; RR-TZF gene subfamily; rice (Oryza sativa L.); off-target effects; inheritable mutagenicity

期刊名称：INTERNATIONAL JOURNAL OF MOLECULAR SCIENCES （ 影响因子：4.9； 五年影响因子：5.7 ）

ISSN： 1661-6596

年卷期： 2025 年 26 卷 3 期

页码：

收录情况： SCI

摘要： The CRISPR/Cas9 system offers a powerful tool for gene editing to enhance rice productivity. In this study, we successfully edited eight RR-TZF genes in japonica rice Nipponbare using CRISPR-Cas9 technology, achieving a high editing efficiency of 73.8%. Sequencing revealed predominantly short insertions or deletions near the PAM sequence, along with multi-base deletions often flanked by identical bases. Off-target analysis identified 5 out of 31 predicted sites, suggesting the potential for off-target effects, which can be mitigated by designing gRNAs with more than three base mismatches. Notably, new mutations emerged in the progeny of several gene-edited mutants, indicating inheritable genetic mutagenicity. Phenotypic analysis of homozygous mutants revealed varied agronomic traits, even within the same gene, highlighting the complexity of gene-editing outcomes. These findings underscore the importance of backcrossing to minimize off-target and inheritable mutagenicity effects, ensuring more accurate trait evaluation. This study offers insights into CRISPR/Cas9 mechanisms and uncertain factors and may inform future strategies for rice improvement, prompting further research into CRISPR/Cas9's precision and long-term impacts.

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