Search-and-replace genome editing without double-strand breaks or donor DNA

Abstract

Most genetic variants that contribute to disease 1 are challenging to correct efficiently and without excess byproducts 2-5. Here we describe prime editing, a versatile and precise genome editing method that directly writes new genetic information into a specified DNA site using a catalytically impaired Cas9 fused to an engineered reverse transcriptase, programmed with a prime editing guide RNA (pegRNA) that both specifies the target site and encodes the desired edit. We performed >175 edits in human cells including targeted insertions, deletions, and all 12 types of point mutations without requiring double-strand breaks or donor DNA templates. We applied prime editing in human cells to correct efficiently and with few byproducts the primary genetic causes of sickle cell disease (requiring a transversion in HBB) and Tay-Sachs disease (requiring a deletion in HEXA), to install a protective transversion in PRNP, and to precisely insert various

CRISPR in livestock: From editing to printing

“…catalytically impaired Cas9 endonuclease fused to a reverse transcriptase allows performing precise targeted insertions, deletions and point mutations without requiring DSBs or donor DNA templates. This approach, known as prime editing, presents lower offtargets activity and fewer byproducts than previous alternatives [20].…”

Self-cutting and integrating CRISPR plasmids (SCIPs) enable targeted genomic integration of genetic payloads for rapid cell engineering

“…While DNA base editors (Gaudelli et al 2017) and their "prime editing" counterparts (Anzalone et al 2019) likely represent the immediate future for performing therapeutic genome editing in settings where very few nucleotides require replacement, there remains a need for simple and expandable tools with which to conduct genome engineering research. In addition to the uses described here (ie.…”

Tumour travel tours – Why circulating cancer cells value company

“…Nevertheless, apart from having become the new standard for generating experimental knock-out models, solid proof has been generated that the method holds the potential to correct serious genetic defaults, such as sickle cell anaemia [31]. Moreover, "prime editing", the in situ directed modification of single nucleotides without the requirement of the cellular DNA repair machinery, hit the headlines recently [32].…”