Frequent and extensive Cas9-directed genomic damage

CRISPR/Cas9 is poised to become the gene editing tool of choice in clinical contexts, but the full spectrum of on-target damage remains unknown. Thus far, exploration of Cas9 induced genetic alterations has been limited to the immediate vicinity of the target site and distal off-target sequences, leading to the conclusion that CRISPR/Cas9 was reasonably specific. Using guide RNAs designed to target benign intronic sites in the mono-allelic PigA and bi-allelic Cd9 genes we detected significant and extensive mutagenesis in the vicinity of the target site.