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Genome-wide profiling of prime editor off-target sites in vitro and in vivo using PE-tag.

Author information

Affiliations

  • 1. RNA Therapeutics Institute, University of Massachusetts Chan Medical School, Worcester, MA, USA.
      Authors
    • Liang SQ 1
    • Chen Z 1
    • Sontheimer EJ 1
    • Xue W 1, 2
    (4 authors)
  • 2. Department of Molecular, Cell and Cancer Biology, University of Massachusetts Chan Medical School, Worcester, MA, USA.
      Authors
    • Liu P 2
    • Ponnienselvan K 2
    • Suresh S 2
    • Zhu LJ 2
    • Xue W 1, 2
    • Wolfe SA 2
    (6 authors)
  • 3. Wyss Institute, Harvard Medical School, Boston, MA, USA.
      Authors
    • Kramme C 3
    • Chatterjee P 3
    (2 authors)

ORCIDs linked to this article

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Nature Methods, 08 May 2023, 20(6):898-907
https://doi.org/10.1038/s41592-023-01859-2 PMID: 37156841 

Abstract 

Prime editors have a broad range of potential research and clinical applications. However, methods to delineate their genome-wide editing activities have generally relied on indirect genome-wide editing assessments or the computational prediction of near-cognate sequences. Here we describe a genome-wide approach for the identification of potential prime editor off-target sites, which we call PE-tag. This method relies on the attachment or insertion of an amplification tag at sites of prime editor activity to allow their identification. PE-tag enables genome-wide profiling of off-target sites in vitro using extracted genomic DNA, in mammalian cell lines and in the adult mouse liver. PE-tag components can be delivered in a variety of formats for off-target site detection. Our studies are consistent with the high specificity previously described for prime editor systems, but we find that off-target editing rates are influenced by prime editing guide RNA design. PE-tag represents an accessible, rapid and sensitive approach for the genome-wide identification of prime editor activity and the evaluation of prime editor safety.

Full text links 

Read article at publisher's site: https://doi.org/10.1038/s41592-023-01859-2

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Funding 

Funders who supported this work.

Foundation for the National Institutes of Health (2)

NCATS NIH HHS (1)

NHLBI NIH HHS (3)

National Institutes of Health