Engineered CRISPR prime editors with compact, untethered reverse transcriptases

Julian Grünewald; Bret R. Miller; Regan N. Szalay; Peter K. Cabeceiras; Christopher J. Woodilla; Eliza Jane B. Holtz; Karl Petri; J. Keith Joung

doi:10.1038/s41587-022-01473-1

Engineered CRISPR prime editors with compact, untethered reverse transcriptases

Julian Grünewald, Bret R. Miller, Regan N. Szalay, Peter K. Cabeceiras, Christopher J. Woodilla, Eliza Jane B. Holtz, Karl Petri, J. Keith Joung

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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.

Original language	English
Pages (from-to)	337-343
Number of pages	7
Journal	Nature Biotechnology
Volume	41
Issue number	3
DOIs	https://doi.org/10.1038/s41587-022-01473-1
State	Published - Mar 2023

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title = "Engineered CRISPR prime editors with compact, untethered reverse transcriptases",

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.",

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AU - Szalay, Regan N.

AU - Cabeceiras, Peter K.

AU - Woodilla, Christopher J.

AU - Holtz, Eliza Jane B.

AU - Petri, Karl

AU - Joung, J. Keith

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AB - 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.

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Engineered CRISPR prime editors with compact, untethered reverse transcriptases

Abstract

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