Keyword

genome editing

5 papers tagged “genome editing”

BiologyNature · Oct 2019 Open access

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

Andrew V. Anzalone, Peyton B. Randolph, Jessie R. Davis and David R. Liu

This paper introduced prime editing, a versatile genome-editing method that uses a Cas9 nickase fused to a reverse transcriptase guided by a prime editing guide RNA (pegRNA) to write new genetic information directly into a target site. Without requiring double-strand breaks or donor DNA, prime editing can install targeted insertions, deletions, and all 12 types of point mutations. The authors demonstrated correction of disease-relevant mutations in human cells with broad targeting flexibility and relatively low off-target activity.

prime editing crispr genome editing reverse transcriptase

BiologyNature · Oct 2017 Open access

Programmable base editing of A•T to G•C in genomic DNA without DNA cleavage

Nicole M. Gaudelli, Alexis C. Komor, Holly A. Rees and David R. Liu

This work developed adenine base editors (ABEs) by evolving a transfer RNA adenosine deaminase to act on DNA, enabling direct conversion of A-T base pairs to G-C in genomic DNA without double-strand breaks. Because no natural DNA adenosine deaminase was available, the authors used directed evolution to create the enzyme, then fused it to Cas9 nickase. ABEs corrected target adenines efficiently and with high product purity and low indel formation in human cells.

base editing adenine base editor crispr genome editing

BiologyNature · Apr 2016 Open access

Programmable editing of a target base in genomic DNA without double-stranded DNA cleavage

Alexis C. Komor, Yongjoo B. Kim, Michael S. Packer, John A. Zuris and David R. Liu

This paper introduced cytosine base editing (CBE), a strategy that fuses a catalytically impaired Cas9 to a cytidine deaminase to directly convert C-G base pairs to T-A in genomic DNA without inducing double-strand breaks or requiring a donor template. The authors showed that within a programmable target window the deaminase converts cytosine to uracil, which is then read as thymine, and that inhibiting base-excision repair markedly improves editing efficiency. The approach achieved precise single-base correction in human and other mammalian cells.

base editing crispr genome editing deaminase

BiologyCell · Sept 2015 Open access

Cpf1 Is a Single RNA-Guided Endonuclease of a Class 2 CRISPR-Cas System

Bernd Zetsche, Jonathan S. Gootenberg and Feng Zhang

This paper characterizes Cpf1 (later named Cas12a) as a single-RNA-guided DNA endonuclease of a class 2 CRISPR-Cas system, expanding the genome-editing toolbox beyond Cas9. Cpf1 requires only a single crRNA (no tracrRNA), recognizes a T-rich PAM, and produces staggered cuts with overhangs. The authors demonstrate Cpf1-mediated genome editing in human cells.

crispr cas12a cpf1 genome editing

BiologyScience · Aug 2012 Open access

A Programmable Dual-RNA–Guided DNA Endonuclease in Adaptive Bacterial Immunity

Martin Jinek, Krzysztof Chylinski, Ines Fonfara, Michael Hauer, Jennifer A. Doudna and Emmanuelle Charpentier

This study demonstrated that the CRISPR-associated protein Cas9 from Streptococcus pyogenes is an RNA-guided DNA endonuclease whose target specificity is determined by a dual-RNA structure formed by a CRISPR RNA (crRNA) base-paired to a trans-activating crRNA (tracrRNA). The authors showed that Cas9 introduces site-specific double-strand breaks in target DNA, with its HNH domain cleaving the complementary strand and its RuvC-like domain cleaving the noncomplementary strand. Critically, they engineered the two guide RNAs into a single chimeric guide RNA that still directed sequence-specific cleavage, establishing the system as a programmable tool for genome editing.

crispr cas9 genome editing molecular biology