Visible traits demonstrate that crispant founder mice can be used for phenotypic assessment

Genes can be knocked out in model organisms by introducing a single guide RNA and Cas9 into one cell zygotes. Recently, the zebrafish and Xenopus communities have employed this method in genetic screening pipelines that assess phenotypes in founders (F0), referred to as “crispants”. In contrast, phenotyping of crispant mice has been avoided as results are believed to be confounded by genetic mosaicism, requiring that only established mouse lines undergo phenotypic assessment. Here, we targeted seven genes associated with visible recessive phenotypes. We observed the expected null phenotype in up to 100% founders per gene. Crucially, we achieved 100% editing efficiency in all but two animals. Genetic mosaicism was common, but did not confound an animal’s phenotype when comprised of mutations that all disrupt the targeted gene. Mosaicism included short in-frame mutations, but these were sufficient to disrupt function of five genes. Several founders were compound heterozygotes carrying a null and a non-null allele (short in-frame mutation or late truncation), enabling functional assessment of the non-null allele to dissect protein function. Our results set the stage for using crispant founders for initial phenotypic assessment in genetic screening, before selecting candidates for further study. This will dramatically reduce animal numbers.