Fig. 1

Schematic approaches to identify off-targeting events. First, genomic DNAs were isolated from knockout pigs or fetuses, generated using the CRISPR/Cas9 system. Second, a PCR reaction was performed to amplify a fragment of DNA flanking off-target region. Third, PCR products were loaded on a gel and Sanger sequenced. If the results were polymorphic, then TOPO cloning followed by Sanger sequencing was performed to uncover the genotype of each allele