Comparative Single-Cell Profiling of CRISPR Knockout and Interference Defines Modality-Specific Strengths in Functional Genomics

Authors

Gupta, N., Sayer, A., Prater, M., Mastrokalou, C., Saeed, K., Company, C., Hart, C., Miragaia, R., Trehan, A., Functional Genomics Centre,, McDermott, U., Strauss, M. E., Ross-Thriepland, D., Walter, D., Kalinka, A.

Abstract

Pooled CRISPR screens coupled with single-cell RNA sequencing enable high-throughput functional interrogation of gene regulatory networks, yet systematic comparisons of CRISPR knockout (CRISPRko) and CRISPR interference (CRISPRi) remain limited. We established a single-cell CRISPRko workflow and benchmarked it against CRISPRi using 87 sgRNAs targeting 29 unfolded protein response genes. CRISPRko generated transcriptional phenotypes are highly concordant with CRISPRi, and induced comparable pathway-level responses. While CRISPRi allows direct assessment of target gene repression, CRISPRko provides an effective complementary approach for complete loss-of-function studies, expanding the toolkit for single-cell functional genomics.

Preprint server: bioRxiv
The authors list and abstract were imported from bioRxiv on 28 Aug 2026.

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