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Enhanced Genome Editing with Cas9 Ribonucleoprotein in Diverse Cells and Organisms
Published on: May 25, 2018
Robust genome and RNA editing via CRISPR nucleases in PiggyBac systems
Yuqian Jiang1,2, Rachel Catherine Hoenisch1, Yun Chang3
1Department of Biomedical Engineering, Pennsylvania State University, University Park, PA, 16802, USA.
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.
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