High-content CRISPR screening
High-content CRISPR screening
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DOI：
10.1038/s43586-021-00093-4
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发表时间：
2022-02-10
期刊：
影响因子：
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通讯作者：
Zhuang, Xiaowei
中科院分区：
文献类型：
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作者：
Bock, Christoph;Datlinger, Paul;Zhuang, Xiaowei
CRISPR筛选是生物学发现的强大来源，可以在广泛的应用和物种中对基因功能进行公正的询问。在汇集的CRISPR筛选中，各种基因编码的扰动被引入细胞池。靶细胞在细胞竞争、药物治疗或病毒感染等生物挑战下增殖。随后，通过对指定每种扰动的向导rna的基于序列的计数来评估微扰诱导的效应。这种筛选的典型结果是对基因进行排序，这些基因对感兴趣的生物挑战具有敏感性或抗性。CRISPR核心技术的适应性使得激活、沉默或以其他方式操纵靶基因成为可能，这有助于CRISPR筛选的广泛应用。此外，单细胞RNA测序和空间成像等高含量读数有助于以前所未有的细节表征筛选的细胞。专用软件工具促进生物信息学分析和提高再现性。CRISPR筛选已经揭示了基础生物学、医学遗传学、癌症研究、免疫学、传染病、微生物学等领域的各种分子机制。本入门介绍了CRISPR筛选的基本和高级概念及其作为生物发现，生物医学研究和药物开发的灵活可靠方法的应用-特别强调高含量方法，可以直接作为筛选的一部分获得详细的生物学见解。
CRISPR screens are a powerful source of biological discovery, enabling the unbiased interrogation of gene function in a wide range of applications and species. In pooled CRISPR screens, various genetically encoded perturbations are introduced into pools of cells. The targeted cells proliferate under a biological challenge such as cell competition, drug treatment or viral infection. Subsequently, the perturbation-induced effects are evaluated by sequencing-based counting of the guide RNAs that specify each perturbation. The typical results of such screens are ranked lists of genes that confer sensitivity or resistance to the biological challenge of interest. Contributing to the broad utility of CRISPR screens, adaptations of the core CRISPR technology make it possible to activate, silence or otherwise manipulate the target genes. Moreover, high-content read-outs such as single-cell RNA sequencing and spatial imaging help characterize screened cells with unprecedented detail. Dedicated software tools facilitate bioinformatic analysis and enhance reproducibility. CRISPR screening has unravelled various molecular mechanisms in basic biology, medical genetics, cancer research, immunology, infectious diseases, microbiology and other fields. This Primer describes the basic and advanced concepts of CRISPR screening and its application as a flexible and reliable method for biological discovery, biomedical research and drug development - with a special emphasis on high-content methods that make it possible to obtain detailed biological insights directly as part of the screen.