Haploid SLCs appeared in the male germline cell induction system in vitro. a Images of the day 21- and day 23-induced cells. Upper panel, bright-field images of the day 21- and day 23-induced cells, arrowheads indicate small round haploid cells. Lower panel, acrosin-DsRed expression profile of these small round cells at day 23. Scale bar, 100 μm. b Real-time PCR analyses of acrosin, Prm1 and Tnp-1 expression levels during the induction process at day 19, day 21, day 23, and day 25. c Flow cytometric analysis of the percentage of haploid cells of day 23-induced cells. d Real-time PCR analyses of acrosin, Prm1, Tnp-1, Haprin, Sycp3, Stra8, Sox2, and Nanog expression level of ESCs, day 23-induced cells and sperms. Data are represented as mean ± SEM from more than three independent experiments, *p < 0.05, **p < 0.01, ***p < 0.001.