Objective To establish an efficient method of genotyping for Lepr ^{db/ +} mouse offsprings by TaqMan probe quantitative fluorescence PCR. Methods Genome DNA was extracted from tails of 228 Lepr ^{db/ +} mouse offsprings. PCR primers and TaqMan probes were designed according to the mutation sites of Lepr gene (rs1801133). Real time PCR assay was applied and SNP loci were typed with SDS software. The genotyping of 2-month old Lepr ^{db/ db} mice was validated by the phenotype and Hardy-Weinberg equilibrium test was performed. Results 228 samples were detected by the established TaqMan fluorescence quantitative PCR assay. 64 mice were of GG genotype, with a genotype frequency of 0.1929. 123 mice were of GT genotype, with a genotype frequency of 0.5395. 41 mice were of TT genotype, with a genotype frequency of 0.2807. Compared with the phenotype typing, the sensitivity of the TaqMan fluorescence quantitative PCR was 97.56% and the specificity was 99.47%. Conclusions TaqMan probe quantitative fluorescence PCR assay is a simple and efficient method, and can be used to detect the genotype of Lepr ^{db/ +} mouse offsprings.