Epigenetics Programme, Babraham Institute, Cambridge CB22 3AT, UK.
Wellcome Trust Sanger Institute, Hinxton, Cambridge CB10 1SA, UK.
STAR Protoc. 2021 Apr 9;2(2):100426. doi: 10.1016/j.xpro.2021.100426. eCollection 2021 Jun 18.
CRISPR/Cas9 screens are a powerful approach to identify key regulators of biological processes. By combining pooled CRISPR/Cas9 screening with single-cell RNA-sequencing readout, individual perturbations can be assessed in parallel both comprehensively and at scale. Importantly, this allows gene function and regulation to be interrogated at a cellular level in an unbiased manner. Here, we present a protocol to perform pooled CRISPR-activation screens in mouse embryonic stem cells using 10× Genomics scRNA-seq as a readout. For complete information on the generation and use of this protocol, please refer to Alda-Catalinas et al. (2020).
CRISPR/Cas9 筛选是鉴定生物过程关键调控因子的有力方法。通过将 pooled CRISPR/Cas9 筛选与单细胞 RNA-seq 读数相结合，可以全面、大规模地同时评估单个扰动。重要的是，这使得可以在无偏倚的方式在细胞水平上研究基因功能和调控。在这里，我们展示了一种在小鼠胚胎干细胞中使用 10× Genomics scRNA-seq 作为读数进行 pooled CRISPR 激活筛选的方案。有关此方案的生成和使用的完整信息，请参阅 Alda-Catalinas 等人（2020 年）。
