Fig. 1.

RT-PCR and flow cytometric analysis. RT-PCR was used to determine the expression of specific spermatogonia and germ cell markers. a Expression of the specific spermatogonia and germ cell-specific markers was detected by RT-PCR of Itgα6 (148 bp), Itgβ1 (3115 bp), Plzf (137 bp), VASA (149 bp), Oct4 (129 bp), and Gfrα-1 (130 bp); genes were expressed in testis tissue (positive control) and spermatogonial cells after culture. Flow cytometric analysis of spermatogonial cells. b The cells without antibody staining considered as negative control, c spermatogonial cells after two-step enzymatic digestion, and d after 2 week in culture. The population of cultured cells were double stained with antibodies against β1-integrin and c-kit. In three experiments, the average percent of the spermatogonial cells (C-KIT⁻, INTEGRIN β₁ ⁺) after two-step enzymatic digestion was 16.06 ± 1.8 %, whereas cultivation enhanced the purity of these cells to 34.9 ± 2.3 %