使用 MAGeCKFlute 对汇集的 CRISPR 遗传筛选进行综合分析。
Integrative analysis of pooled CRISPR genetic screens using MAGeCKFlute.
机构信息
Shanghai Key Laboratory of Tuberculosis, Clinical Translational Research Center, Shanghai Pulmonary Hospital, School of Life Sciences and Technology, Tongji University, Shanghai, China.
Department of Geriatrics, Shanghai General Hospital, Shanghai, China.
出版信息
Nat Protoc. 2019 Mar;14(3):756-780. doi: 10.1038/s41596-018-0113-7. Epub 2019 Feb 1.
Genome-wide screening using CRISPR coupled with nuclease Cas9 (CRISPR-Cas9) is a powerful technology for the systematic evaluation of gene function. Statistically principled analysis is needed for the accurate identification of gene hits and associated pathways. Here, we describe how to perform computational analysis of CRISPR screens using the MAGeCKFlute pipeline. MAGeCKFlute combines the MAGeCK and MAGeCK-VISPR algorithms and incorporates additional downstream analysis functionalities. MAGeCKFlute is distinguished from other currently available tools by its comprehensive pipeline, which contains a series of functions for analyzing CRISPR screen data. This protocol explains how to use MAGeCKFlute to perform quality control (QC), normalization, batch effect removal, copy-number bias correction, gene hit identification and downstream functional enrichment analysis for CRISPR screens. We also describe gene identification and data analysis in CRISPR screens involving drug treatment. Completing the entire MAGeCKFlute pipeline requires ~3 h on a desktop computer running Linux or Mac OS with R support.
利用 CRISPR 与核酸酶 Cas9（CRISPR-Cas9）进行全基因组筛选是一种用于系统评估基因功能的强大技术。需要进行基于统计学原理的分析，才能准确识别基因命中和相关途径。在这里，我们描述了如何使用 MAGeCKFlute 管道进行 CRISPR 筛选的计算分析。MAGeCKFlute 结合了 MAGeCK 和 MAGeCK-VISPR 算法，并纳入了其他下游分析功能。MAGeCKFlute 与其他当前可用的工具的区别在于其全面的管道，其中包含一系列用于分析 CRISPR 筛选数据的功能。本协议解释了如何使用 MAGeCKFlute 来执行 CRISPR 筛选的质量控制 (QC)、归一化、批次效应去除、拷贝数偏差校正、基因命中识别和下游功能富集分析。我们还描述了涉及药物处理的 CRISPR 筛选中的基因识别和数据分析。在配备 R 支持的 Linux 或 Mac OS 桌面计算机上，完成整个 MAGeCKFlute 管道大约需要 3 小时。
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