C1 single-cell RNA-seq of Adult ID4-EGFP mouse spermatoognia

To reveal distinct transcriptomes associated with spermatogonial stem cell renewal vs. initiation of differentiation, single-cell transcriptomes from Adult 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. Four replicate preparations of Adult mouse ID4-EGFP+ spermatogonia were used for this study. Data are from a total of 300 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.为揭示与精原干细胞（spermatogonial stem cell, SSC）自我更新及分化起始相关的差异化转录组特征，本研究对成年ID4-增强绿色荧光蛋白阳性（ID4-EGFP+）精原细胞的单细胞转录组进行细分，获得经移植实验验证后具有不同分化命运的细胞亚群：其中ID4-EGFP强阳性（ID4-EGFPbright）细胞高度富集精原干细胞（SSCs），而ID4-EGFP弱阳性（ID4-EGFPdim）细胞则富集于精原祖细胞。本研究采用Fluidigm C1单细胞捕获仪器捕获单个精原细胞，利用SMART-Seq2技术开展单细胞RNA测序。本研究共使用4批成年小鼠ID4-EGFP+精原细胞的重复制备样本，最终纳入总计300个细胞的测序数据。为便于组间比较，本研究根据EGFP表观荧光强度将所有细胞划分为4个四分位组（Q1为EGFP强阳性，Q4为EGFP弱阳性，Q2与Q3为EGFP荧光强度中等的细胞群）。