The escape of CRISPR-mediated gene editing in Zymomonas mobilis

文献类型： 外文期刊

第一作者： Chen, Mao

作者： Chen, Mao;Huang, Yuhuan;Zheng, Yudi;Wu, Bo;He, Mingxiong;Chen, Mao;Huang, Yuhuan;Zheng, Yudi

作者机构：

关键词： CRISPR; Zymomonas mobilis; knockout; plasmid curing; gene escape; literature review

期刊名称：FEMS MICROBIOLOGY LETTERS （ 影响因子：2.1； 五年影响因子：2.8 ）

ISSN： 0378-1097

年卷期： 2023 年 370 卷

页码：

收录情况： SCI

摘要： Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR) systems have been widely applied for gene or genome editing. Adequate checking is important to screen mutants after CRISPR-mediated editing events. Here, we report gene escape cases after the knockout by Type I-F native CRISPR system in Zymomonas mobilis. Through amplifying both the gene of interest and its flanking homologous arms, followed by curing the editing plasmid, we found different destinies for gene-editing events. Some genes were readily knocked out and followed by the easy plasmid curing. In some other cases, however, the editing plasmid was difficult to remove from the cell, or the deleted genes were transferred into the editing plasmid. For example, the targeted region of fur can be integrated into the editing plasmid after the knockout, resulting in a spurious editing event. We supposed that the transfer of the gene may be attributed to bacterial insertion sequences. Searching for literatures on the gene knockout using CRISPR in bacteria reveals that the escape event is likely underestimated due to inadequate validation in other microbes. Hence, several strategies are proposed to enhance gene knockout and plasmid curing. CRISPR-mediated gene editing in Zymomonas mobilis revealed the different destinies including gene escape from various events.

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