Use of CRISPR/Cas9 to Induce Targeted Mutagenesis during Porcine Embryogenesis

Introducing the CRISPR/Cas9 system into early embryos can introduce targeted modifications during embryogenesis. Previous reports suggest that the approach can be used to generate pigs with targeted modifications without having to generate genetically engineered founder pigs for breeding. We have used this approach to disrupt various genes during embryogenesis in the pig. A total of 22 target genes were selected and CRISPR/Cas9 was introduced into early pig embryos to disrupt each target gene. Genotype results demonstrate that the CRISPR/Cas9 system could introduce targeted modifications on all target genes in all embryos examined. Until now, in vivo efficacy of the approach has been confirmed in 9 genes. In addition, we have successfully attempted to modify up to three genes simultaneously (100%) demonstrating its ability to multiplex. Our results indicate that the technology can be successfully used to introduce targeted modifications in the pig.