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

By Nicole M. Gaudelli, Alexis C. Komor, H. Rees, M. Packer, A. Badran, D. Bryson, David R. Liu. 2017

5,379 citations recorded by the source corpus.

The spontaneous deamination of cytosine is a major source of transitions from C•G to T•A base pairs, which account for half of known pathogenic point mutations in humans. The ability to efficiently convert targeted A•T base pairs to G•C could therefore advance the study and treatment of genetic diseases. The deamination of adenine yields inosine, which is treated as guanine by polymerases, but no enzymes are known to deaminate adenine in DNA. Here we describe adenine base editors (ABEs) that mediate the conversion of A•T to G•C in genomic DNA. We evolved a transfer RNA adenosine deaminase to operate on DNA when fused to a catalytically impaired CRISPR–Cas9 mutant. Extensive directed evolution and protein engineering resulted in seventh-generation ABEs that convert targeted A•T base pairs efficiently to G•C (approximately 50% efficiency in human cells) with high product purity (typically at least 99.9%) and low rates of indels (typically no more than 0.1%). ABEs introduce point mutations more efficiently and cleanly, and with less off-target genome modification, than a current Cas9 nuclease-based method, and can install disease-correcting or disease-suppressing mutations in human cells. Together with previous base editors, ABEs enable the direct, programmable introduction of all four transition mutations without double-stranded DNA cleavage.

Subfields: Molecular Biology

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Citations in the corpus

Evolved Cas9 variants with broad PAM compatibility and high DNA specificity (2018, neutral): “2a and Extended Data Figs 4–6) in HEK293T cells.”
MGnify: the microbiome analysis resource in 2020 (2019, background): “For example, at present, all 96 individual studies that make up the Earth Microbiome Project (18) are grouped together under a single Super Study, as are MGnify analyses for 8 distinct Tara Oceans (19) datasets.”
DRAM for distilling microbial metabolism to automate the curation of microbiome function (2020, background): “Scores highlighted in red are considered high (1, 2) or medium (3) confidence and thus the putative AMGs are also represented in the Product.”
What is Speciation Genomics? The roles of ecology, gene flow, and genomic architecture in the formation of species (2018, background): “To summarize, advances in CRISPR techniques are developing at astonishing rates, including methods to directly convert single bases without requiring the formation of double-stranded breaks, using a catalytically impaired CRISPR/Cas9 mutant base editing (Nishida et al., 2016; Gaudelli et al., 2017).”
A meta-analysis of the effect of organic and mineral fertilizers on soil microbial diversity (2020, methodology): “For example, meta-analysis of observational data suggest that biological richness (measured by DNA sequences) is greatest at neutral pH and at a mean temperature of 10 °C (Thompson et al., 2017).”
Gene therapy comes of age (2018, positive): “Correction of mutations at a single-base level via Cas9based targeting of “base editors”has recently been reported (61, 62).”