使用和基因组编辑对遗传变异的功能评估的协议
Sofia Obolenski1, Rebeca Olvera-León2, Dijue Sun2
1Experimental Cancer Genetics, Wellcome Sanger Institute, Wellcome Trust Genome Campus, Hinxton, CB10 1SA Cambridge, UK; Department of Dermatology, Leiden University Medical Centre, 2333 ZA Leiden, the Netherlands.
STAR protocols
|April 8, 2025
概括
和基因组编辑 (SGE) 提供了一种使用CRISPR-Cas9和同源导向修复 (HDR) 的多重核酸修改方法. 本协议详细介绍了HAP1-A5细胞中变异评估的SGE,使功能遗传分析成为可能.
科学领域:
- 分子生物学分子生物学
- 基因组学就是基因组学.
- 生物技术是生物技术.
背景情况:
- 和基因组编辑 (SGE) 是一种用于多重核酸修饰的强大技术.
- 克里斯普尔-Cas9和同质导向修复 (HDR) 是SGE的关键组成部分.
- 对遗传变异的功能分析需要保存原生基因组背景的方法.
研究的目的:
- 提出基于SGE的变种评估的详细协议.
- 为了使HAP1-A5细胞中的遗传变异能够进行功能分析.
- 概述设计变体库,sgRNA和PCR.原始剂的步骤.
主要方法:
- 设计变体库,单导向RNA (sgRNA) 和PCR的寡核酸原始剂.
- 用于SGE选的样品准备.
- 细胞查和下一代测序 (NGS) 库的准备. 细胞查和下一代测序 (NGS) 库的准备.
主要成果:
- 该协议允许在特定的基因组部位进行详尽的核酸修改.
- 多重编辑允许对遗传变异进行全面的功能分析.
- 该方法保留了评估变异的原生基因组背景.
结论:
- 该协议为基于SGE的变种评估提供了一个强大的框架.
- 描述的方法有助于基因变异的功能基因组学.
- 基因基因学是理解遗传变异及其功能影响的宝贵工具.
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