Genome-wide target specificities of CRISPR RNA-guided programmable deaminases
- Authors
- Kim D.; Lim K.; Kim S.-T.; Yoon S.-H.; Kim K.; Ryu S.-M.; Kim J.-S.
- Issue Date
- May-2017
- Publisher
- Nature Publishing Group
- Citation
- Nature Biotechnology, v.35, no.5, pp 475 - 480
- Pages
- 6
- Indexed
- SCI
SCIE
SCOPUS
- Journal Title
- Nature Biotechnology
- Volume
- 35
- Number
- 5
- Start Page
- 475
- End Page
- 480
- URI
- https://scholarworks.korea.ac.kr/kumedicine/handle/2020.sw.kumedicine/5661
- DOI
- 10.1038/nbt.3852
- ISSN
- 1087-0156
1546-1696
- Abstract
- Cas9-linked deaminases, also called base editors, enable targeted mutation of single nucleotides in eukaryotic genomes. However, their off-target activity is largely unknown. Here we modify digested-genome sequencing (Digenome-seq) to assess the specificity of a programmable deaminase composed of a Cas9 nickase (nCas9) and the deaminase APOBEC1 in the human genome. Genomic DNA is treated with the base editor and a mixture of DNA-modifying enzymes in vitro to produce DNA double-strand breaks (DSBs) at uracil-containing sites. Off-target sites are then computationally identified from whole genome sequencing data. Testing seven different single guide RNAs (sgRNAs), we find that the rAPOBEC1-nCas9 base editor is highly specific, inducing cytosine-to-uracil conversions at only 18 ± 9 sites in the human genome for each sgRNA. Digenome-seq is sensitive enough to capture off-target sites with a substitution frequency of 0.1%. Notably, off-target sites of the base editors are often different from those of Cas9 alone, calling for independent assessment of their genome-wide specificities. © Nature America, Inc., part of Springer Nature. All rights reserved.
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Collections - 1. Basic Science > Department of Physiology > 1. Journal Articles
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