美国普林斯顿大学Britt Adamson、加州大学旧金山分校Jonathan S. Weissman等研究人员合作取得一项新成果。他们开发出直接向导RNA（sgRNA）捕获和靶向测序的组合式单细胞CRISPR筛选技术。2020年3月30日，《自然—生物技术》在线发表了这一成果。
Title: Combinatorial single-cell CRISPR screens by direct guide RNA capture and targeted sequencing
Author: Joseph M. Replogle, Thomas M. Norman, Albert Xu, Jeffrey A. Hussmann, Jin Chen, J. Zachery Cogan, Elliott J. Meer, Jessica M. Terry, Daniel P. Riordan, Niranjan Srinivas, Ian T. Fiddes, Joseph G. Arthur, Luigi J. Alvarado, Katherine A. Pfeiffer, Tarjei S. Mikkelsen, Jonathan S. Weissman, Britt Adamson
Abstract: Single-cell CRISPR screens enable the exploration of mammalian gene function and genetic regulatory networks. However, use of this technology has been limited by reliance on indirect indexing of single-guide RNAs (sgRNAs). Here we present direct-capture Perturb-seq, a versatile screening approach in which expressed sgRNAs are sequenced alongside single-cell transcriptomes. Direct-capture Perturb-seq enables detection of multiple distinct sgRNA sequences from individual cells and thus allows pooled single-cell CRISPR screens to be easily paired with combinatorial perturbation libraries that contain dual-guide expression vectors. We demonstrate the utility of this approach for high-throughput investigations of genetic interactions and, leveraging this ability, dissect epistatic interactions between cholesterol biogenesis and DNA repair. Using direct capture Perturb-seq, we also show that targeting individual genes with multiple sgRNAs per cell improves efficacy of CRISPR interference and activation, facilitating the use of compact, highly active CRISPR libraries for single-cell screens. Last, we show that hybridization-based target enrichment permits sensitive, specific sequencing of informative transcripts from single-cell RNA-seq experiments.