Complete loss of proc results in late onset lethality. (A) proc is duplicated in tandem, with near complete identify at the nucleotide and amino acid sequence level. Two sgRNAs (arrowheads) targeting identical sequences in both copies of exons 4 and 10 produced a 17.3-kb gene deletion resulting in nearly complete ablation of both proc copies. The locus is displayed in 5′ to 3′ orientation, but the actual genomic orientation is in reverse, and thus the copies are named according to the latter nomenclature. (B) proc1 is the predominant expressed copy of proc. Qualitative reverse transcription PCR with primers that match identical sequences in both copies reveals 2 bands that correspond to proc1 and proc2 predicted sizes. (C) DNA gel electrophoresis using a single primer pair that amplifies both copies showed the truncated genomic proc PCR product (0.3 kb) after CRISPR-mediated deletion. (D) qPCR (quantitative PCR) data of total proc expression revealed a 63% decrease in the heterozygous mutant embryos, and 99.8% decrease in homozygous mutants, (P < .0001 by t testing). (E) Survival curves indicated significant mortality in proc^−/− mutants by 1 year of age using log-rank (Mantel-Cox) testing (P < .0001). Chr, chromosome.