Firstly, we analyzed the rare damaging variants in 30 hypospadias risk associated genes (Chen et al., 2019), including SHH, GLI1, GLI2, GLI3, FGF8, FGF10, FGFR2, BMP7, WT1, DGKK, HSD3B1, HSD17B3, SRD5A2, AR, ESR1, ESR2, BMP4, HOXA4, HOXB6, MAP3K1, CHD7, NR5A1, MAMLD1, HSD3B2, CYP11A1, AKR1C3, ATF3, BNC2, SP1, SP7. Among all these detected rare variants in the 30 associated genes of the three hypospadias cases, four rare damaging variants were identified from three patients. Here, GLI1 is linked to hypospadias.