Engineered CRISPR prime editors with compact, untethered reverse transcriptases

Nat Biotechnol. 2023 Mar;41(3):337-343. doi: 10.1038/s41587-022-01473-1. Epub 2022 Sep 26.

Abstract

The CRISPR prime editor PE2 consists of a Streptococcus pyogenes Cas9 nickase (nSpCas9) fused at its C-terminus to a Moloney murine leukemia virus reverse transcriptase (MMLV-RT). Here we show that separated nSpCas9 and MMLV-RT proteins function as efficiently as intact PE2 in human cells. We use this Split-PE system to rapidly identify and engineer more compact prime editor architectures that also broaden the types of RTs used for prime editing.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • CRISPR-Cas Systems* / genetics
  • Deoxyribonuclease I / genetics
  • Gene Editing* / methods
  • Humans
  • Mice
  • Moloney murine leukemia virus* / genetics
  • RNA-Directed DNA Polymerase* / genetics
  • Streptococcus pyogenes* / genetics

Substances

  • RNA-Directed DNA Polymerase
  • Deoxyribonuclease I