GSM8901065: Pool 1, Epithelium, Estrus; Mus musculus; RNA-Seq
The uterine horns were dissected and divided into approximately four segments, which were then placed in a 1% trypsin solution. Following digestion for 45 minutes at 37°C, the uterine epithelium was carefully separated using a mouth pipette and fine tweezers. The isolated epithelium was further digested with 5 mg/ml collagenase and mechanically disrupted into a single-cell suspension using a syringe plunger. After staining with DAPI, live cells were sorted using a cell sorter prior to library construction. Library was construed by following the manufactor's manual (10x Genomics Chromium Single Cell 3' Gene Expression v4 protocol). Briefly, cells, master mix, gel beads, and partitioning oil onto the Chromium Chip and run on the Chromium Controller to generate Gel Bead Emulsions (GEMs). GEMs was incubated on PCR thermocycler for reverse transcription. The cDNA was enzymatically fragmented, end repaired, ligated with sequencing adapater, and amplified with sample-specific index primers. Library was purtified using SPRIselect beads, quantified with Qubit, and assesssed for size distribution using TapeStation before sequencing.