AbstractThis is the first work using gonads from undifferentiated, genetically‐sexed Siberian sturgeon describing expression changes in genes related to steroid synthesis and female and male sex differentiation. One factor identified as relevant for ovarian differentiation was the gene coding for the enzyme Hsd17b1, which converts estrone into estradiol‐17β. hsd17b1 was highly activated in female gonads at 2.5 months of age, around the onset of sex differentiation, preceding activation of two other genes involved in estrogen production (cyp19a1 and foxl2). hsd17b1 was also strongly repressed in males. Two known foxl2 paralogs are found in Siberian sturgeon—foxl2 and foxl2l—but only foxl2 appeared to be associated with ovarian differentiation. With regard to the male pathway, neither 11‐oxygenated androgens nor classic male genes (amh, dmrt1, sox9, and dhh) were found to be involved in male sex differentiation, leaving open the question of which genes participate in early male gonad development in this ancient fish. Taken together, these results indicate an estrogen‐dependence of female sex differentiation and 11‐oxygenated androgen‐independence of male sex differentiation.