DNA修复背景与目标序列的相互作用可预测地偏向了Cas9产生的突变
Ananth Pallaseni1, Elin Madli Peets1, Gareth Girling1
1Wellcome Sanger Institute, Wellcome Genome Campus, Hinxton, United Kingdom.
bioRxiv : the preprint server for biology
|July 10, 2023
概括
CRISPR/Cas9基因编辑结果取决于DNA修复途径. 这项研究揭示了突变生成中的特定基因作用,通过了解DNA修复机制,可以更精确地控制CRISPR/Cas9编辑.
科学领域:
- 分子生物学分子生物学
- 遗传学 是一个遗传学.
- 生物技术是生物技术.
背景情况:
- 通过CRISPR/Cas9基因编辑,引入了双链断裂 (DSB).
- DSB修复途径和侧翼序列会影响突变性结果.
- 修复基因和突变类型之间的相互作用仍未得到充分探索.
研究的目的:
- 在各种修复环境中全面绘制CRISPR/Cas9 DSBs的突变性结果.
- 为了确定影响不同突变类型的特定DNA修复基因.
- 开发CRISPR/Cas9编辑结果的预测模型.
主要方法:
- 使用Cas9 DSBs在小鼠胚胎干细胞中的2838个合成点产生了83,680个独特的突变.
- 在缺少18个关键DNA修复基因的情况下评估突变频率.
- 分类突变类型并将它们与特定的基因淘汰相关联.
主要成果:
- 在特定的1bp插入中确定了DNA-PKcs (Prkdc) 和Polμ (Polm) 的作用.
- 证明了NBN和Polθ (Polq) 在删除形成中的可变参与.
- 发现了依赖Ku80 (Xrcc5) 和NBN.的单向删除.
结论:
- 无偏见的突变分类为新的DSB修复机制提供了洞察力.
- 基于修复基因功能的预测模型优于CRISPR/Cas9结果预测的当前标准.
- 对DNA修复基因功能的更好理解有助于精确调制CRISPR/Cas9诱导的突变.
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