Fig. 1. Characterization of mouse Gabra6⁺ GCPs by fluorescence-activated cell sorting (FACS).

A Schematic model of the cultivation of mouse GCPs purified via discontinuous density gradient centrifugation from the cerebella (postnatal days 5–7) of mice [15, 16] in the presence of NAC (30 μM) for 7 days [10, 11], followed by characterization and FACS-based sorting using an anti-Gabra6 antibody. B FACS-based sorting to enrich the subfraction of Gabra6⁺ GCPs from WT and Jdp2KO mice. The left panel depicts a representative dot plot of FSC and SSC. The region was gated in dot plots and Gabra6⁺ cells were sorted by FACS analysis. The histogram was gated on a peak, to identify the percentage of GCPs that expressed Gabra6. C Staining of Gabra6⁺ GCPs from WT and Jdp2 KO mice with an anti-Atoh1 antibody. The nuclei of PGCs were stained with DAPI. Relative ratio of Atoh1-positive cells among Gabra6⁺ GCPs from WT and Jdp2 KO cells. Scale bars, 50 μm. D Relative values of the Oct4, Sox2, Nonog, Klf4, and c-Myc mRNAs between WT and Jdp2 KO Gabra6⁺ GCPs. The primer sequences used for qPCR are listed in Supplementary Table 2. The levels of the WT cells were taken as 1.0. E Immunostaining of Gabra6⁺ GCPs from WT and Jdp2 KO mice for Nrf2, p21^Cip1, and AhR. The GC nuclei were stained with DAPI. The levels of the WT cells were taken as 1.0. Scale bars, 20 μm. F Comparative expression of the Lgr5 proteins, as assessed by Western blotting (left panel; the quantitation of each protein is shown on the right panel). The relative value was normalized to that of β-actin and presented as a ratio. Uncropped raw data was shown in Fig. S5. B–F: n = 5; *P < 0.05, **P < 0.01; ***P < 0.001.