Generation of dual-gRNA library for combinatorial CRISPR screening of synthetic lethal gene pairs.
Generation of dual-gRNA library for combinatorial CRISPR screening of synthetic lethal gene pairs.
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DOI：
10.1016/j.xpro.2022.101556
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发表时间：
2022-09-16
期刊：
影响因子：
--
通讯作者：
Li, Lang
中科院分区：
文献类型：
--
作者：
Tang, Shan;Wu, Xue;Liu, Jinghui;Zhang, Qiongsi;Wang, Xinyi;Shao, Shuai;Gokbag, Birkan;Fan, Kunjie;Liu, Xiaoqi;Li, Fuhai;Cheng, Lijun;Li, Lang
组合CRISPR筛选可用于研究合成致死性（SL）基因对。在这里，我们详细介绍了双gRNA文库构建的步骤，并引入了两个骨架，LentiGuide_DKO和LentiCRISPR_DKO。我们描述了用22 Rv 1-Cas9和SaOS 2-Cas9细胞进行体外筛选，然后进行测序和数据分析的步骤。通过引入两个主链，我们优化了文库构建过程，促进了标准的对端测序，并提供了在具有或不具有Cas9表达修饰的细胞上筛选的选择。通过引入两个主链优化文库构建过程体外筛选基因-基因相互作用w/wo Cas9从基因选择到数据分析的完整筛选过程出版商说明：进行任何实验方案都需要遵守当地机构的实验室安全和伦理指南。组合CRISPR筛选可用于研究合成致死性（SL）基因对。在这里，我们详细介绍了双gRNA文库构建的步骤，并引入了两个骨架，LentiGuide_DKO和LentiCRISPR_DKO。我们描述了用22 Rv 1-Cas9和SaOS 2-Cas9细胞进行体外筛选，然后进行测序和数据分析的步骤。通过引入两个主链，我们优化了文库构建过程，促进了标准的对端测序，并提供了在具有或不具有Cas9表达修饰的细胞上筛选的选择。
Combinatorial CRISPR screening is useful for investigating synthetic lethality (SL) gene pairs. Here, we detail the steps for dual-gRNA library construction, with the introduction of two backbones, LentiGuide_DKO and LentiCRISPR_DKO. We describe steps for in vitro screening with 22Rv1-Cas9 and SaOS2-Cas9 cells followed by sequencing and data analysis. By introducing two backbones, we optimized the library construction process, facilitated standard pair-end sequencing, and provided options of screening on cells with or without modification of Cas9 expression. Optimized library construction process by introducing two backbones In vitro screening gene-gene interaction w/wo Cas9 Full screening process from gene selection to data analysis Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics. Combinatorial CRISPR screening is useful for investigating synthetic lethality (SL) gene pairs. Here, we detail the steps for dual-gRNA library construction, with the introduction of two backbones, LentiGuide_DKO and LentiCRISPR_DKO. We describe steps for in vitro screening with 22Rv1-Cas9 and SaOS2-Cas9 cells followed by sequencing and data analysis. By introducing two backbones, we optimized the library construction process, facilitated standard pair-end sequencing, and provided options of screening on cells with or without modification of Cas9 expression.
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