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Robust genome and RNA editing via CRISPR nucleases in PiggyBac systems.

ABSTRACT: CRISPR/Cas-mediated genome editing in human pluripotent stem cells (hPSCs) offers unprecedented opportunities for developing in vitro disease modeling, drug screening and cell-based therapies. To efficiently deliver the CRISPR components, here we developed two all-in-one vectors containing Cas9/gRNA and inducible Cas13d/gRNA cassettes for robust genome editing and RNA interference respectively. These vectors utilized the PiggyBac transposon system, which allows stable expression of CRISPR components in hPSCs. The Cas9 vector PB-CRISPR exhibited high efficiency (up to 99%) of inducing gene knockout in both protein-coding genes and long non-coding RNAs. The other inducible Cas13d vector achieved extremely high efficiency in RNA knockdown (98% knockdown for CD90) with optimized gRNA designs. Taken together, our PiggyBac CRISPR vectors can serve as powerful toolkits for studying gene functions in hPSCs.

SUBMITTER: Jiang Y

PROVIDER: S-EPMC8964983 | biostudies-literature | 2022 Aug

REPOSITORIES: biostudies-literature

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Robust genome and RNA editing via CRISPR nucleases in PiggyBac systems.

Jiang Yuqian Y Hoenisch Rachel Catherine RC Chang Yun Y Bao Xiaoping X Cameron Craig E CE Lian Xiaojun Lance XL

Bioactive materials 20220207

CRISPR/Cas-mediated genome editing in human pluripotent stem cells (hPSCs) offers unprecedented opportunities for developing <i>in vitro</i> disease modeling, drug screening and cell-based therapies. To efficiently deliver the CRISPR components, here we developed two all-in-one vectors containing Cas9/gRNA and inducible Cas13d/gRNA cassettes for robust genome editing and RNA interference respectively. These vectors utilized the PiggyBac transposon system, which allows stable expression of CRISPR ...[more]

PMID: 35386818

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