GSM9080373: GF pancreas, AP, rep3; Mus musculus; RNA-Seq
Fresh pancreas was minced (~0.5 mm³) in DMEM + 10 % FBS and digested at 37 °C for 60 min in DMEM containing 0.25 % trypsin and 1 mg ml⁻¹ collagenase I/II/IV with gentle agitation. The digest was filtered through a 100 µm strainer and centrifuged (300 g, 5 min). Red blood cells were removed with MACS RBC lysis buffer (4 °C, 10 min). Cells were washed, resuspended in PBS + 0.04 % BSA, and viability was assessed on a Luna counter; suspensions were adjusted to 700–1 200 cells µl⁻¹ for library preparation. Single-cell libraries were generated using Chromium Next GEM Single Cell 3′ Reagent Kits v3.1 (10x Genomics, PN 1000268) following the manufacturer's instructions, targeting 8 000 cells per sample. GEM generation, reverse transcription, cDNA amplification and indexing PCR were performed on a Chromium Controller. Finished libraries were quantified (Qubit dsDNA HS) and quality-checked (Agilent 2100). Sequencing was carried out on an Illumina NovaSeq 6000 (paired-end 2 × 150 bp).