Pa5 31177

Western blotting (WB) was estimated according to a previous article [27 (link)]. The antibodies used in this study are listed as follows: anti-eIF2γ (1 : 500; PA5-31177, Thermo Fisher Scientific, MA, USA), anti-PCNA (1 : 500; BM0104, Boster, Wuhan, China), anti-Cyclin D (1 : 500; WL01435a, Wanlei, Shenyang, China), anti-GAPDH (1 : 2000; AC002, ABclonal, Wuhan, China), anti-ZBTB16 (1 : 500; No. D222893, BBI Life, Shanghai, China), anti-pERK (1 : 2000; #4370, CST, Boston, USA), and anti-ERK (1 : 2000; #9194, CST, Boston, USA). The results were detected using a Bio-Rad imaging system (Bio-Rad, Hercules, CA, USA) and quantified using ImageJ (V1.48d).

Immunofluorescence staining of testes and SSCs was conducted as previously reported [27 (link)]. The primary antibodies used in this study are listed as follows: rabbit anti-eIF2γ (1 : 200; PA5-31177, Thermo Fisher Scientific, MA, USA), rabbit anti-DDX4 (1 : 200; ab13840, Abcam, Cambridge, UK), mouse anti-ZBTB16 (1 : 200; sc-28319, Santa Cruz Biotechnology, CA, USA), rabbit anti-STRA8 (1 : 200; ab49602, Abcam, Cambridge, UK), mouse anti-GFRa1 (1 : 200; sc-271546, Santa Cruz Biotechnology, CA, USA), rabbit anti-SOX9 (1 : 200; ab185230, Abcam, Cambridge, UK), and rabbit anti-StAR (1 : 100; bs-20388R, Bioss, Beijing, China). Secondary antibodies are as follows: Alexa Fluor 488-goat anti-rabbit IgG (1 : 400; ZF-0511, ZSGB-BIO, Beijing, China) and Alexa Fluor 568-goat anti-mouse (1 : 400; ZF-0513, ZSGB-BIO, Beijing, China).