Differences between immunodeficient mice generated by classical gene targeting and CRISPR/Cas9-mediated gene knockout

Immunodeficient mice are widely used for pre-clinical studies to understand various human diseases. Here, we report the generation of four immunodeficient mouse models using CRISPR/Cas9 system without inserting any foreign gene sequences such as Neo R cassettes and their characterization. By eliminating any possible effects of adding a Neo R cassette, our mouse models may allow us to better elucidate the in vivo functions of each gene. Our FVB- Rag2 −/− , B6- Rag2 −/− , and BALB/c- Prkdc −/− mice showed phenotypes similar to those of the earlier immunodeficient mouse models, including a lack of mature B cells and T cells and an increase in the number of CD45 + DX-5 + natural killer cells. However, B6- Il2rg −/− mice had a unique phenotype, with a lack of mature B cells, increased number of T cells, and decreased number of natural killer cells. Additionally, serum immunoglobulin levels in all four immunodeficient mouse models were significantly reduced when compared to those in wild-type mice with the exception of IgM in B6- Il2r g −/− mice. These results indicate that our immunodeficient mouse models are a robust tool for in vivo studies of the immune system and will provide new insights into the variation in phenotypic outcomes resulting from different gene-targeting methodologies.