CRISPR-based genome perturbation provides a new avenue to conveniently change DNA sequences, transcription, and epigenetic modifications in genetic screens. However, it remains challenging to assay the complex molecular readouts after perturbation at high resolution and at scale. By introducing an A/G mixed capture sequence into the gRNA scaffold, we demonstrate that gRNA transcripts could be directly reverse transcribed by poly (dT) primer together with the endogenous mRNA, followed by high-content molecular phenotyping in scRNA-seq (Direct-seq). With this method, the CRISPR perturbation and its transcriptional readouts can be profiled together in a streamlined workflow.

中文翻译：

---

Direct-seq：用于 CRISPR 筛选中简化 scRNA-seq 的编程 gRNA 支架


基于 CRISPR 的基因组扰动提供了一种在基因筛选中方便地改变 DNA 序列、转录和表观遗传修饰的新途径。然而，在高分辨率和大规模扰动后分析复杂的分子读数仍然具有挑战性。通过将 A/G 混合捕获序列引入 gRNA 支架中，我们证明 gRNA 转录物可以通过聚 (dT) 引物与内源 mRNA 一起直接逆转录，然后在 scRNA-seq（Direct-seq）中进行高内涵分子表型分析。序列）。通过这种方法，可以在简化的工作流程中一起分析 CRISPR 扰动及其转录读数。