We demonstrate CRISPR-Cas9-mediated spermatogenic cell-specific disruption of Scp3 gene in testes in one step.Female transgenic mice were generated by pronuclear injection of a plasmid containing Hspa2 promoter driven Cas9 and constitutive expressed sgRNA targeting to Scp3 gene.Male progeny were infertile due to developmental arrest of spermatogenic cells.Our study establishes a one-step strategy to analyze gene function in adult tissues by a temporal-spatial pattern.