A more efficient method to generate null mutants using Hprt-Cre with floxed alleles

Abstract

Purpose The generation of nonviable homozygous null mouse embryos from heterozygote null/+ breedings can be highly resource consuming, with only 25% of the embryos in the litter being null mutants. We hypothesized that (1) we could double the number of homozygous null mouse embryos in a litter without reducing litter size using Hypoxanthine-guanine phosphoribosyltransferase-Cre (Hprt)-Cre (which is active in the female germ line at the time of fertilization), and (2) these homozygous null mutants would be identical to mutants generated through traditional null/+ breedings. Methods To test this hypothesis, we used a conditional allele Fgfr2IIIb flox . This allele when recombined is identical to the Fgfr2IIIb null allele. An F1 generation of Fgfr2IIIb rec/+ ; Hprt Cre/+ females was created by mating Fgfr2IIIb +/+ ; Hprt cre /cre females to a Fgfr2IIIb flox/flox male. The F1 females were then mated to a Fgfr2IIIb flox/flox male. F2 embryos were genotyped, and the morphology and histology of the lungs, intestine, limbs, and brain were analyzed. Results The Hprt-Cre mating strategy results in 51% of pups being genotypic homozygous null embryos (85/166) vs 23% for the standard null/+ approach (38/167). These embryos did not express the Fgfr2IIIb transcript and were phenotypically identical to null embryos generated through standard null/+ breedings. Conclusions The Hprt-Cre mating strategy increases the number of homozygous mutant embryos in a litter without decreasing litter size. Embryos generated through this approach are phenotypically identical to those from standard heterozygous breedings. We recommend this approach to investigators using a model system that relies on the generation of homozygous null embryos.