Modified pegRNAs mitigate scaffold-derived prime editing by-products

Panagiotis Antoniou, Louis Dacquay, Hanna Mårtensson, Katja Madeyski-Bengtson, Anna-Lena Loyd, Anna Shiriaeva, Euan Gordon, Salman Mustfa, George Thom, Pei-Pei Hsieh, Saša Šviković, Mike Firth, Nina Akrap, Marcello Maresca () and Martin Peterka ()
Additional contact information
Panagiotis Antoniou: AstraZeneca
Louis Dacquay: AstraZeneca
Hanna Mårtensson: AstraZeneca
Katja Madeyski-Bengtson: AstraZeneca
Anna-Lena Loyd: AstraZeneca
Anna Shiriaeva: AstraZeneca
Euan Gordon: AstraZeneca
Salman Mustfa: AstraZeneca
George Thom: AstraZeneca
Pei-Pei Hsieh: AstraZeneca
Saša Šviković: AstraZeneca
Mike Firth: AstraZeneca
Nina Akrap: AstraZeneca
Marcello Maresca: AstraZeneca
Martin Peterka: AstraZeneca

Nature Communications, 2025, vol. 16, issue 1, 1-11

Abstract: Abstract Prime editors (PEs) employ reverse transcriptase (RT) to install genomic edits using a template within the prime editing guide RNA (pegRNA). RT creates a 3’ genomic flap containing the intended edit. However, reverse transcription can continue beyond the template, incorporating the pegRNA scaffold sequence into the 3’ flap. These scaffold-derived by-products can be installed alongside the intended edit, reducing prime editing precision. Here, we develop a method that prevents RT from accessing the scaffold, thereby mitigating such by-products. We demonstrate that an internal abasic spacer or 2’-O-methylation within the pegRNAs terminates RT at the end of the template. This prevents scaffold-derived sequences from being incorporated into the target locus. We benchmark these pegRNAs in different cell types and demonstrate that they can be used with processive PEs such as PE6d or PE**. Our findings provide a simple approach to mitigate a common prime editing by-product and improve prime editing precision.

Date: 2025
References: View references in EconPapers View complete reference list from CitEc
Citations:

Downloads: (external link)
https://www.nature.com/articles/s41467-025-58653-1 Abstract (text/html)

Related works:
This item may be available elsewhere in EconPapers: Search for items with the same title.

Export reference: BibTeX RIS (EndNote, ProCite, RefMan) HTML/Text

Persistent link: https://EconPapers.repec.org/RePEc:nat:natcom:v:16:y:2025:i:1:d:10.1038_s41467-025-58653-1

Ordering information: This journal article can be ordered from
https://www.nature.com/ncomms/

DOI: 10.1038/s41467-025-58653-1

Access Statistics for this article

Nature Communications is currently edited by Nathalie Le Bot, Enda Bergin and Fiona Gillespie

More articles in Nature Communications from Nature
Bibliographic data for series maintained by Sonal Shukla () and Springer Nature Abstracting and Indexing ().