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移除TREX1活动可以增强CRISPR-Cas9介导的同源重组组合

Mehmet E Karasu¹, Eléonore Toufektchan², Yanyang Chen²

¹Department of Biology, Institute of Molecular Health Sciences, Swiss Federal Institute of Technology (ETH) Zurich, Zurich, Switzerland.

Nature biotechnology

|August 12, 2024

概括

TREX1外核酶活动阻碍了CRISPR-Cas9基因编辑效率. 抑制TREX1或使用受保护的DNA模板可显著增强不同细胞类型的同质导向修复 (HDR).

科学领域:

分子生物学分子生物学
遗传学是一个遗传学.
基因编辑技术的技术

背景情况:

在精确的基因组工程中,CRISPR-Cas9介导的同质导向修复 (HDR) 是至关重要的.
实现高HDR效率仍然是一个重大挑战,特别是在某些细胞类型中,如那些具有DNA修复缺陷的细胞类型.

研究的目的:

为了确定限制CRISPR-Cas9 HDR效率的遗传因素.
在具有挑战性的细胞环境中发现改善HDR的策略.

主要方法:

在Fanconi贫血患者的淋巴细胞细胞系中进行全基因组查.
评估外核酶TREX1对HDR效率的影响.
评估TREX1淘汰和化学保护的单链DNA (ssDNA) 模板用于HDR增强.

主要成果:

在使用ssDNA或线性双链DNA修复模板时,TREX1被确定为CRISPR-Cas9 HDR的关键抑制剂.
在各种细胞系和初级细胞中观察到的高TREX1表达与HDR效率差相关.
通过TREX1淘汰赛或使用TREX1抗性ssDNA模板,可以将HDR效率提高两到八倍.

结论:

TREX1活动解释了不同细胞类型的可变CRISPR-Cas9 HDR效率.
针对TREX1或使用受保护的DNA模板提供了一种可行的策略,以促进TREX1表达细胞中的基因编辑结果.

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