Three novel NR5A1 mutations. (A) Schematic overview of the positions of the mutations and electropherograms. (B) RT-qPCR showed a lower NR5A1 expression in the maternal grandfather of the index patient (I:1), and in the mother of the index patient (II:2). We did not include the index case in this experiment as no fresh blood could be collected. Two negative control samples (NC) without the mutation were included for comparison. To exclude technical variations, expression of the reference genes GADPH, HMBS and TBP were also measured, showing stable expression in all patients. (C) Pedigrees for the patients with a NR5A1 mutation. The genotype of the analysed individuals is shown under their symbol. Full black squares indicate affected males with hypospadias, partially black circles indicate females with POF and circles with a black dot correspond with asymptomatic carrier females.