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• W3214328950 abstract "Genome-wide unbiased identification of double-stranded breaks enabled by sequencing (GUIDE-seq) is a sensitive, unbiased, genome-wide method for defining the activity of genome-editing nucleases in living cells. GUIDE-seq is based on the principle of efficient integration of an end-protected double-stranded oligodeoxynucleotide tag into sites of nuclease-induced DNA double-stranded breaks, followed by amplification of tag-containing genomic DNA molecules and high-throughput sequencing. Here we describe a detailed GUIDE-seq protocol including cell transfection, library preparation, sequencing and bioinformatic analysis. The entire protocol including cell culture can be completed in 9 d. Once tag-integrated genomic DNA is isolated, library preparation, sequencing and analysis can be performed in 3 d. The result is a genome-wide catalog of off-target sites ranked by nuclease activity as measured by GUIDE-seq read counts. GUIDE-seq is one of the most sensitive cell-based methods for defining genome-wide off-target activity and has been broadly adopted for research and therapeutic use. GUIDE-seq (genome-wide unbiased identification of double-stranded breaks enabled by sequencing) is a sensitive, unbiased, genome-wide method for defining the specificity of genome-editing nucleases in living cells." @default.
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• W3214328950 date "2021-11-12" @default.
• W3214328950 modified "2023-10-16" @default.
• W3214328950 title "Defining genome-wide CRISPR–Cas genome-editing nuclease activity with GUIDE-seq" @default.
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• W3214328950 doi "https://doi.org/10.1038/s41596-021-00626-x" @default.
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