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Efficient Genome Editing of Mice by CRISPR Electroporation of Zygotes
Published on: December 16, 2022
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Efficient engineering of human and mouse primary cells using peptide-assisted genome editing
Zhen Zhang1,2, Amy E Baxter3,4, Diqiu Ren1,5,6
1Epigenetics Institute, University of Pennsylvania, Philadelphia, PA, USA.
Nature Biotechnology
|April 24, 2023
Summary
A new Peptide-Assisted Genome Editing (PAGE) system enables rapid and safe CRISPR genome editing in primary cells. This method achieves high editing efficiencies with minimal toxicity, overcoming key delivery challenges.
Area of Science:
- Molecular Biology
- Gene Editing Technologies
- Cellular Biology
Background:
- Efficient delivery of CRISPR genome editing tools into primary cells is a significant hurdle.
- Existing methods like electroporation can cause cellular toxicity and transcriptional changes.
Purpose of the Study:
- To develop a novel, efficient, and well-tolerated CRISPR delivery system for primary cells.
- To establish a rapid and robust genome editing platform with minimal off-target effects.
Main Methods:
- Development of the engineered Peptide-Assisted Genome Editing (PAGE) CRISPR-Cas system.
- Utilizing cell-penetrating Cas9/Cas12a and endosomal escape peptides for delivery.
- Incubation of primary cells with PAGE components for 30 minutes.
Main Results:
- Achieved robust single and multiplex genome editing in various primary cells.
- Demonstrated high editing efficiencies (upwards of 98%) in human and mouse T cells and hematopoietic progenitor cells.
- Observed low cellular toxicity and no significant transcriptional perturbation compared to electroporation.
Conclusions:
- The PAGE system offers a broadly generalizable platform for next-generation genome engineering in primary cells.
- PAGE provides a simple, efficient, and safe method for CRISPR delivery, overcoming previous limitations.
