Fig. 6: Avian ES cells are competent to give rise to germ cells.

a, RT–qPCR analysis of the expression of germ cell markers in chicken ES cells (day 0) and during EB differentiation. Data represent mean ± s.d. of three biological replicates. b, Immunofluorescence staining of DAZL in day-3 EBs formed from chicken or goose ES cells. n = 3 independent experiments. Scale bars, 10 μm. c, GFP fluorescence images showing the testes dissected from an E15 high-grade chimera generated by injecting GFP-labeled chicken ES cells into the subgerminal cavity of an EGK.X chicken embryo. Scale bars, 50 μm. d, Images showing GFP and DAZL double staining of tissue sections from E9 and E15 testes dissected from chimeras generated by injecting GFP-labeled chicken ES cells into the subgerminal cavity of EGK.X chicken embryos. Scale bars, 10 μm. e, Images showing GFP and SSEA-1 double staining of tissue sections from an E15 gonad dissected from a chimera generated by injecting GFP-labeled chicken ES cells into the subgerminal cavity of an EGK.X chicken embryo. Scale bars, 10 μm. f, Immunostaining of E22 testis sections from a chimera generated by injecting GFP-labeled chicken ES cells into the subgerminal cavity of an EGK.X-stage chicken embryo. The right panel shows a zoom-in view of the boxed area in the left panel. Arrows indicate GFP and DAZL double-positive cells; arrowheads indicate GFP-negative/DAZL-positive cells. Scale bars, left, 200 µm, right, 50 µm. g, Images showing GFP/P63/SOX9 triple staining of tissue sections from E21 testes dissected from chimeras generated by injecting GFP-labeled chicken ES cells into the subgerminal cavity of EGK.X chicken embryos. Arrows indicate GFP⁺/P63⁺/SOX9⁻ germ cells (GC) in the top row and GFP⁺/P63⁻/SOX9⁺ Sertoli cells (SC) in the bottom row. Scale bars, 20 μm. Chimeras shown in c–g are from one representative experiment out of five performed.