Gonadal differentiation associated with estrogen-induced female sex determination was examined in a turtle with temperature-dependent sex determination, and was compared to ovarian differentiation at a femaleproducing temperature. Freshly laid eggs of the redeared slider. Trachemys scripta. were incubated at a male-producing temperature (26°C) and were experimentally manipulated at one of three embryonic stages: stage 15, 17, or 20 (i.e. early. midway, or late in the temperature-sensitive and estrogen-sensitive periods). At those developmental stages. groups of eggs were either (1) treated with a control solution (95% ethanol) and placed back at the male-producing temperature. (2) treated with 10 μg of estradiol-17β and placed back at the male-producing temperature. or (3) shifted to a female-producing temperature (31°C). Additionally. a control group of freshly laid eggs was continually incubated at 31°C throughout embryonic development. To examine morphological events occurring after the treatments, a subset of embryos from each group was examined at the time of the treatment and at 1–2 stage intervals following the treatments. The results indicate that estradiol-17β as well as female-producing temperature may ensure female sex determination by facilitating medullary cord regression. Further. the results reveal a chronology of differentiation in which medullary cord regression temporally precedes cortical proliferation. © 1993, International Society of Differentiation. All rights reserved.