在CRISPR-Cas9诱导的断裂中,基因组的重复性和调节性元素和供体序列的普遍整合
Chongwei Bi1, Baolei Yuan1, Yingzi Zhang1
1Bioscience Program, Biological and Environmental Science and Engineering Division, King Abdullah University of Science and Technology (KAUST), Thuwal, 23955-6900, Kingdom of Saudi Arabia.
Communications biology
|January 20, 2025
概括
克里斯普尔-Cas9基因编辑可以导致大型的意外插入 (LgIns),通常涉及移动遗传元素. 化DNA捐赠者可显著降低这些基因毒性风险,而不会影响编辑效率.
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:
- 克里斯普尔-Cas9基因编辑被广泛使用,但它具有诸如大型插入 (LgIns) 等基因毒性风险.
- 方法上的局限性阻碍了对这些意外的大插入的研究.
研究的目的:
- 综合调查和报告由CRISPR-Cas9.9诱导的意外大插入 (LgIns).
- 为了确定这些插入的性质及其起源.
- 评估减轻这些基因毒性风险的策略.
主要方法:
- 对CRISPR-Cas9编辑结果在各种基因位置,Cas9疗法和捐赠者类型的分析.
- 插入序列的表征,包括可逆转移的元素和宿主基因组片段.
- 评估3D基因组组织和DNA修复机制.
- 对化双链DNA (dsDNA) 捐赠者的评估,以减少LgIns.
主要成果:
- 通过不同的CRISPR-Cas9方法持续诱导意想不到的LgIns,经常包含可逆转移的元素和宿主基因组序列.
- 有证据表明,LgIns是由通过修复途径随机获取DNA片段引起的.
- 观察到全长和合的dsDNA捐赠者的显著意外整合.
- 酸化的dsDNA捐赠者将LgIns和删除减少了近一半,同时保持了同质导向修复 (HDR) 的效率.
结论:
- CRISPR-Cas9编辑带来了大量的,被低估的意外大插入的风险.
- 了解LgIns的机制对于安全的基因编辑应用至关重要.
- 化dDNA供体提供了一个有希望的策略,以提高CRISPR-Cas9技术的安全性.
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