GSM5509343: mouse_D5all_10X; Mus musculus; RNA-Seq
Cells were isolated from ligated right carotid arteries of VSMC lineage-labeled animals (Myh11-EYFP-Ki67/RFP) 5 days after surgery. EYFP+ VSMCs were mixed with EYFP- cells, and 10,000 cells were loaded onto the Chromium system (10x Genomics). Single cell preparation: Carotid arteries were dissected free of perivascular adipose tissue, cut open longitudinally and the endothelium was removed by gentle scraping with a cotton bud. Carotid arteries were then pre-digested in DMEM (Gibco) supplemented with 1 mg/mL Collagenase IV (Invitrogen) and 1 U/mL Elastase (Worthington) for 10 minutes at 37˚C to allow removal of the adventitial layer. The medial layer was transferred to fresh DMEM supplemented with 2.5 mg/mL Collagenase IV (Invitrogen) and 2.5 U/mL Elastase (Worthington) and digested to a single cell suspension at 37 ˚C. A total of 10,000 cells were loaded onto a Chromium system (10x Genomics) and amplified cDNA libraries generated using the 3' Gene Expression v3.0 kit (10x Genomics) were sequenced using 150 cycle paired end protocol (NovaSeq, Illumina).