GSM8635733: Pancreatic islets, Spopfl/fl females, 12-weeks-old; Mus musculus; RNA-Seq
Islets were isolated by collagenase digestion of the dissected pancreas as previously described (Good et al., 2019). After three rounds of hand-picking under a light microscope, islets were allowed to recover for 3 days in RPMI 1640 containing 10 mM glucose.washed with PBS, and dispersed to single cells with 0.25% trypsin at 37°C with gentle pipetting. 40% volume of FBS was added to the solution and the cell suspension was filtered through a 40 μm cell strainer to facilitate further breakup of cells (Corning). Debris and dead cells were then removed from the cell suspension using Dead Cell and Debris Removal kits (Miltenyi Biotec). Cells were counted and resuspended in 1x PBS with 0.04% BSA for library preparation For library preparations, samples were loaded onto a Chromium 10X Genomics instrument to generate single-cell barcoded droplets according to the manufacturer's instructions with 10X single-cell 3' v3.1 Next GEM chemistry. Libraries were sequenced on a Novaseq 6000 instrument (Illumina) with the v1.5 SP 100 cycles reagent kit with parameters 28x10x10x90.