Fig5d Sperm staining of caudal epididymal sperms.

Raw image dataset for sperm staining of caudal epididymal sperms from Wdr62 genetrap and control mice. An incision was made to the caudal end of isolated epididymus in DMEM (Gibco). This was followed by 15 min incubation at 37°C to allow spermatozoa to swim out. Sperms were centrifugated at 150g for 5 min to remove DMEM. Sperms were fixed in 95% ethanol for 15 min and 10μL was smeared onto a glass slide and air-dried for 2 hrs. Slides were kept at 4°C overnight. The next day, sperm stain was performed using the Papanicolaou method as described in https://doi.org/10.1093/eshremonographs/2002.2.13, with 3 min incubation in Harris hematoxylin (Sigma), 2 min in orange G6 (OG6 0.5% solution in 95% ethanol, phosphotungstic acid 0.015g/100ml) and 5 min in EA50 (0.5% lightgreen SF yellowish CI 42095, 0.5% bismark brown CI 21000, 0.5% eosin Y CI 45380, 0.2g phosphotungstic acid and 2 drops of lithium carbonate (0.154g in 10ml dH2O)).本原始图像数据集源自Wdr62基因陷阱（genetrap）小鼠与对照小鼠的附睾尾部精子染色实验，具体实验操作流程如下：将分离得到的附睾尾部置于DMEM培养基（Gibco）中，并于附睾尾部制作切口；随后于37℃下孵育15分钟，使精子游出；将精子以150g离心5分钟以去除DMEM培养基；使用95%乙醇将精子固定15分钟，取10μL滴涂于载玻片上，室温晾干2小时；将载玻片置于4℃条件下过夜保存。次日，按照https://doi.org/10.1093/eshremonographs/2002.2.13中描述的帕帕尼科拉乌（Papanicolaou）染色法进行精子染色：先将样本置于哈里斯苏木精（Harris hematoxylin，Sigma）中孵育3分钟，再于橙黄G6（OG6：95%乙醇配制的0.5%浓度溶液，含磷钨酸0.015g/100ml）中孵育2分钟，最后于EA50染液（成分为：0.5%亮绿SF黄 CI 42095、0.5%俾斯麦棕 CI 21000、0.5%伊红Y CI 45380、0.2g磷钨酸，以及2滴碳酸锂溶液（0.154g溶于10ml去离子水））中孵育5分钟。