GSM8252292: Ovary, AMH, 4 hour; Mus musculus; RNA-Seq
One ovary from each animal was collected in cold PBS, and the ovaries from each group (N = 5 per group) were cut into small pieces and placed in 2 mL of DMEM (10% FBS, 1% penicillin/streptomycin), supplemented with 21 U/mL of Papain (Worthington, LK003178) and 1 mg/mL of collagenase A (Sigma Aldrich, 10103578001), for 30 minutes at 37°C. After digestion, the cells were subjected to further mechanical digestion using the gentleMACS™ Dissociator (Miltenyi Biotec). Single cells were isolated by straining the digested tissue suspension through a 70-micron filter. The cells were then counted to assess cell viability and to obtain 10,000 cells from each group for library preparation. Samples were processed for scRNAseq according to the manufacturer's instructions using the Chromium Next GEM Single Cell 3ʹ Reagent Kits v3.1 (Dual Index) (10× Genomics) and Chromium Controller (10× Genomics). Single-cell libraries were sequenced using the NovaSeq S4 (Illumina) by the Bauer Core Facility at Harvard University.