用基本编辑屏幕对人类变体进行大规模并行评估
Ruth E Hanna1, Mudra Hegde1, Christian R Fagre2
1Genetic Perturbation Platform, Broad Institute, Cambridge, MA 02142, USA.
Cell
|February 19, 2021
概括
这项研究引入了可扩展的CRISPR-Cas9基编辑选方法,以快速确定遗传变异的功能影响. 这种方法可以准确地识别引起疾病的突变,并揭示耐药性机制.
科学领域:
- 遗传学
- 分子生物学
- 生物技术
背景情况:
- 鉴定单核酸变体 (SNV) 对于了解疾病遗传学至关重要.
- 目前用于变种表征的技术在可扩展性和吞吐量方面有限.
研究的目的:
- 使用CRISPR-Cas9基编辑器开发和验证可扩展的选方法,以测量哺乳动物细胞内源的SNV.
- 展示基因编辑器屏幕在检测药物变异相互作用和DNA损伤修复基因变异的有用性.
主要方法:
- 在聚合屏幕中使用CRISPR-Cas9细胞基编辑器对数千个SNV进行测试.
- 在正面和负面选择屏幕上对基准编辑器的性能进行了比较.
- 创建了一个单向导RNAs (sgRNAs) 的库,针对3,584个基因中的52,034个ClinVar变体.
主要成果:
- 在识别BRCA1和BRCA2已知的功能丧失突变方面取得了高精度.
- 鉴定了对BH3模拟剂和PARP抑制剂产生药物敏感性或耐药性的点突变.
- 在细胞应激条件下发现了DNA损伤修复基因的功能丧失变异.
结论:
- CRISPR-Cas9基编辑器屏幕提供了一个可扩展和高效的基因变异功能平台.
- 这种方法加快了对疾病和药物反应的遗传变异后果的理解.
- 该方法广泛适用于高通量功能基因组学和变异解释.
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