C1 single-cell RNA-seq of immature (P6) ID4-EGFP mouse spermatoognia

To reveal distinct transcriptomes associated with spermatogonial stem cell renewal vs. initiation of differentiation, single-cell transcriptomes from P6 ID4-EGFP+ spermatogonia were subdivided into subpopulations that displayed distinct fates when assayed by transplantation, with ID4-EGFPbright cells highly enriched for SSCs, and ID4-EGFPdim cells enriched for progenitors. We used the Fluidigm C1 instrument to capture individual spermatogonia for SMART-Seq2 single-cell RNA-seq. Five replicate preparations of mouse P6 ID4-EGFP+ spermatogonia were used for this study. Data are from a total of 278 cells. Cells were binned into quartiles according to EGFP epifluorescence intensity (Q1 = EGFP-bright, Q4=EGFP-dim, Q2 and Q3 have intermediate EGFP fluorescence) to facilitate comparisons.为阐明与精原干细胞自我更新及分化启动相关的独特转录组特征，本研究针对出生后6天（P6）的ID4-EGFP阳性精原细胞开展单细胞转录组分析，并将其细分为经移植实验检测后呈现不同细胞命运的亚群：其中ID4-EGFP亮阳性（ID4-EGFPbright）细胞高度富集精原干细胞（spermatogonial stem cell, SSCs），而ID4-EGFP暗阳性（ID4-EGFPdim）细胞则富集于祖细胞群体。本研究采用Fluidigm C1系统捕获单个精原细胞，依托SMART-Seq2技术完成单细胞RNA测序。本次实验共使用5批重复制备的P6龄小鼠ID4-EGFP阳性精原细胞样本，最终纳入278个细胞的测序数据。为便于组间比较，我们依据EGFP落射荧光强度将细胞划分为四分位数分组：Q1对应EGFP强阳性（亮）组，Q4对应EGFP弱阳性（暗）组，Q2与Q3为EGFP荧光强度处于中间水平的组别。