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A versatile, high-efficiency platform for CRISPR-based gene activation

ABSTRACT: CRISPR-mediated transcriptional activation (CRISPRa) is a powerful technology for inducing gene expression from endogenous loci with exciting applications in high throughput gain-of-function genomic screens and the engineering of cell-based models. However, current strategies for generating potent, stable, CRISPRa-competent cell lines present limitations for the broad utility of this approach. Here, we provide a high-efficiency, self-selecting CRISPRa enrichment strategy, which combined with piggyBac transposon technology enables rapid production of CRISPRa-ready cell populations compatible with a variety of downstream assays. We complement this with an optimized guide RNA scaffold that significantly enhances CRISPRa functionality. Finally, we describe a synthetic guide RNA tool set that enables transient, population-wide gene activation when used with the self-selecting CRISPRa system. Taken together, this versatile platform greatly enhances the potential for CRISPRa across a wide variety of cellular contexts. The generation of CRISPR-mediated transcriptional activation (CRISPRa)-competent cell lines pose significant technical challenges. Here the authors report a platform for production of CRISPRa-ready cell populations which they combine with optimised expressed and synthetic gRNA scaffolds to enhance functionality.

SUBMITTER: Heidersbach A

PROVIDER: S-EPMC9938141 | biostudies-literature | 2023 Jan

REPOSITORIES: biostudies-literature

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