Zhang Z, Baxter AE, Ren D, Qin K, Chen Z, Collins SM, Huang H, Komar CA, Bailer PF, Parker JB, Blobel GA, Kohli RM, Wherry EJ, Berger SL, Shi J. Efficient engineering of human and mouse primary cells using peptide-assisted genome editing. Nat Biotechnol. 2024 Feb;42(2):305-315. doi: 10.1038/s41587-023-01756-1. Epub 2023 Apr 24. PMID: 37095348.


Simple, efficient and well-tolerated delivery of CRISPR genome editing systems into primary cells remains a major challenge. Here we describe an engineered Peptide-Assisted Genome Editing (PAGE) CRISPR-Cas system for rapid and robust editing of primary cells with minimal toxicity. The PAGE system requires only a 30-min incubation with a cell-penetrating Cas9 or Cas12a and a cell-penetrating endosomal escape peptide to achieve robust single and multiplex genome editing. Unlike electroporation-based methods, PAGE gene editing has low cellular toxicity and shows no significant transcriptional perturbation. We demonstrate rapid and efficient editing of primary cells, including human and mouse T cells, as well as human hematopoietic progenitor cells, with editing efficiencies upwards of 98%. PAGE provides a broadly generalizable platform for next-generation genome engineering in primary cells.