Robust genome and RNA editing via CRISPR nucleases in PiggyBac systems

Yuqian Jiang; Rachel Catherine Hoenisch; Yun Chang; Xiaoping Bao; Craig E. Cameron; Xiaojun Lance Lian

Bioactive Materials (Aug 2022)

Robust genome and RNA editing via CRISPR nucleases in PiggyBac systems

Yuqian Jiang,
Rachel Catherine Hoenisch,
Yun Chang,
Xiaoping Bao,
Craig E. Cameron,
Xiaojun Lance Lian

Affiliations

Yuqian Jiang: Department of Biomedical Engineering, Pennsylvania State University, University Park, PA, 16802, USA; Huck Institutes of the Life Sciences, Pennsylvania State University, University Park, PA, 16802, USA
Rachel Catherine Hoenisch: Department of Biomedical Engineering, Pennsylvania State University, University Park, PA, 16802, USA
Yun Chang: Davidson School of Chemical Engineering, Purdue University, West Lafayette, IN, 47907, USA
Xiaoping Bao: Davidson School of Chemical Engineering, Purdue University, West Lafayette, IN, 47907, USA; Corresponding author.
Craig E. Cameron: Department of Microbiology and Immunology, University of North Carolina School of Medicine, Chapel Hill, NC, 27599, USA
Xiaojun Lance Lian: Department of Biomedical Engineering, Pennsylvania State University, University Park, PA, 16802, USA; Huck Institutes of the Life Sciences, Pennsylvania State University, University Park, PA, 16802, USA; Department of Biology, Pennsylvania State University, University Park, PA, 16802, USA; Corresponding author. Department of Biomedical Engineering, Pennsylvania State University, University Park, PA, 16802, USA.

Journal volume & issue: Vol. 14
pp. 313 – 320

Abstract

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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.

Published in Bioactive Materials

ISSN: 2452-199X (Online)
Publisher: KeAi Communications Co., Ltd.
Country of publisher: China
LCC subjects: Technology: Electrical engineering. Electronics. Nuclear engineering: Materials of engineering and construction. Mechanics of materials; Science: Biology (General)
Website: https://www.keaipublishing.com/en/journals/bioactive-materials/

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