Programmable site-specific DNA double-strand breaks via PNA-assisted prokaryotic Argonautes

Research output: Contribution to journalArticlepeer-review


Programmable site-specific nucleases promise to unlock myriad applications in basic biology research, biotechnology and gene therapy. Gene-editing systems have revolutionized our ability to engineer genomes across diverse eukaryotic species. However, key challenges, including delivery, specificity and targeting organellar genomes, pose barriers to translational applications. Here, we use peptide nucleic acids (PNAs) to facilitate precise DNA strand invasion and unwinding, enabling prokaryotic Argonaute (pAgo) proteins to specifically bind displaced single-stranded DNA and introduce site-specific double-strand breaks (DSBs) independent of the target sequence. We named this technology PNA-assisted pAgo editing (PNP editing) and determined key parameters for designing PNP editors to efficiently generate programable site-specific DSBs. Our design allows the simultaneous use of multiple PNP editors to generate multiple site-specific DSBs, thereby informing design considerations for potential in vitro and in vivo applications, including genome editing.
Original languageEnglish (US)
StatePublished - Aug 10 2023

Bibliographical note

KAUST Repository Item: Exported on 2023-09-05
Acknowledged KAUST grant number(s): BAS/1/1035-01-01
Acknowledgements: This work was supported by BAS/1/1035-01-01 baseline and KAUST Smart Health Initiative (KSHI) funding to M.M. We would like to thank members of the Laboratory for Genome Engineering and Synthetic Biology for their discussions and help. We would like to thank Lixin Ma and Fei Wang, State Key Laboratory of Biocatalysis and Enzyme Engineering, School of Life Sciences, Hubei University, Wuhan, Hubei, China, for providing KmAgo expression plasmid.

ASJC Scopus subject areas

  • Genetics


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