AAV9 induces the Sycp1 promoter to drive transgene expression in germ cells in vivo. (A) Schematic illustration of the process used to screen for germ-cell-specific gene promoters that drive RFP reporter gene expression in distinct germ cell types in vivo; four independent germ-cell-specific gene promoter (i.e., Stra8, Hspa2, Sycp1, and Pgk2) were individually fused to the RFP reporter gene, packaged into AAV9, and then microinjected into the seminiferous tubule of testis of 3-week-old mice. (B) Stereomicroscopic fluorescence imaging of representative testes microinjected with AAV9-Stra8-RFP, AAV9-Hspa2-RFP, AAV9-Sycp1-RFP, or AAV9-Pgk2-RFP. Scale bar: 1 mm. (C) Immunostaining of testicular sections from testes received microinjection with AAV9-Stra8-RFP、AAV9-Hspa2-RFP、AAV9-Sycp1-RFP and AAV9-Pgk2-RFP for WT1 (green) and RFP (red). Nuclei were stained with DAPI (blue). Scale bar: 50 μm. (D) Representative stereomicroscopic fluorescent images of testes on days 5, 7, 14, 21, and 28 after microinjection of AAV9-Sycp1-RFP. Scale bar: 1 mm. (E) Immunostaining of testicular sections on days 5, 7, 14, 21, and 28 after microinjection of AAV9-Sycp1-RFP for WT1 (yellow), DDX4 (green), and RFP (red). Nuclei were stained with DAPI (blue). Scale bar: 50 μm.