GSM5195927: DDC; Mus musculus; RNA-Seq
The aorta and heart of three mice in each group were digested with enzyme solution [2mg/ml collagenase I (Worthington Biochemical Corporation, LS004196), 60 U/ml DNase I (Roche Diagnostics, 1010459001)] at 37°C for 30-45 minutes with shaking. The cell suspension was filtered through a 40 μm strainer and enriched for endothelial cells using CD31 MicroBeads (Miltenyi Biotec, 130097418) according to the manufacturer's instructions. The CD31-enriched single cell suspension was washed with WB and stained with Viable dye eFluor 450 (Thermo Fisher Scientific, 50-112-8817). Doublets were gated out prior to FACS sorting, and the viable cells were sorted into a collecting tube with DMEM containing 20% FBS. The cell suspensions were loaded on a 10x Genomics Chromium instrument to generate single cell gel beads in emulsion (GEMs). Libraries were prepared using the Chromium Single cell 3' Reagent Kits v3.1 (CG000204): Chromium Single Cell 3' Library & Gel Bead Kit v3.1 (PN-1000128), Chromium Single Cell 3' Chip Kit v2 (PN-1000127) and Single Index Kit Set A (PN-1000213). Libraries were sequenced on an Illumina NovaSeq 6000 using paired-end sequencing with approximately >90% sequencing saturation.