Unbiased detection of off-target cleavage by CRISPR-Cas9 and TALENs using integrase-defective lentiviral vectors

• DOI: 10.1038/nbt.3127
• Publication Date: 2015-01-19T16:45:15Z
• Authors: Xiaoling Wang; Yebo Wang; Xiwei Wu; Jinhui Wang; Yingjia Wang; Zhaojun Qiu; Tammy Chang; He Huang; Ren-Jang Lin; Jiing-Kuan Yee
• Keywords: 0303 health sciences; 03 medical and health sciences; HEK293 Cells; Integrases; Genome, Human; Genetic Vectors; Lentivirus; High-Throughput Nucleotide Sequencing; Humans; RNA Editing; CRISPR-Cas Systems

The utility of CRISPR-Cas9 and TALENs for genome editing may be compromised by their off-target activity. We show that integrase-defective lentiviral vectors (IDLVs) can detect such off-target cleavage with a frequency as low as 1%. In the case of Cas9, we find frequent off-target sites with a one-base bulge or up to 13 mismatches between the single guide RNA (sgRNA) and its genomic target, which refines sgRNA design.