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Identifying genome-wide off-target sites of CRISPR RNA-guided nucleases and deaminases with Digenome-seq.
Nat Protoc
February 2021
Center for Genome Engineering, Institute for Basic Science (IBS), Seoul, Republic of Korea.
Digested genome sequencing (Digenome-seq) is a highly sensitive, easy-to-carry-out, cell-free method for experimentally identifying genome-wide off-target sites of programmable nucleases and deaminases (also known as base editors). Genomic DNA is digested in vitro using clustered regularly interspaced short palindromic repeats ribonucleoproteins (RNPs; plus DNA-modifying enzymes to cleave both strands of DNA at sites containing deaminated base products, in the case of base editors) and subjected to whole-genome sequencing (WGS) with a typical sequencing depth of 30×. A web-based program is available to map in vitro cleavage sites corresponding to on- and off-target sites.
View Article and Find Full Text PDFDigenome-seq web tool for profiling CRISPR specificity.
Nat Methods
May 2017
Department of Chemistry, Hanyang University, Seoul, South Korea.
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