Gonadal differentiation in marsupials takes place after birth. Germ cell development is initiated in the fetus, and the gonadal ridge is formed but is not sexually dimorphic morphologically by the day of birth. The testicular cords form by day 2 post partum, and the ovarian cortex and medulla several days later, by day 8.  Because testicular and ovarian differentiation occurs over a prolonged time period of several weeks, gonadal differentiation in the tammar wallaby gives an opportunity to investigate the network of genes involved in testicular and ovarian differentiation in a way that is not possible in species like mice where the molecular changes are completed in just a few days.

We investigated SRY, SOX9, FGF9, AMH, WNT4, FOXL2, RSPO1 and FST in the tammar wallaby. Tammar SRY mRNA was localised in pre-Sertoli and Sertoli cells, not only during the gonadal determination window, but also after sexual differentiation, as in the human but in contrast to the mouse. Tammar SOX9 and AMH were detected in both testes and ovaries from before birth, but became sexually dimorphic after sexual differentiation. FGF9 and WNT4 had contrasting mRNA expression patterns in tammar and mouse, suggesting their conserved and antagonistic roles in gonadal differentiation. FOXL2, FST and RSPO1 were expressed in the somatic cells of both testis and ovary before birth, but during the period of gonadal sex determination were strongly expressed in the ovary, but not in the testis, suggesting these are critical genes in ovarian development. Thus, even in our marsupial model, with its extended period of sexual differentiation, there is a high degree of conservation of the sex determining pathway, demonstrating that the molecular control and genes involved in gonadal differentiation has been tightly constrained during 160 million years since the evolution of therian mammals.