GSM9634857: IVCs, DVT; Mus musculus; RNA-Seq
Mouse infrarenal IVCs were collected and sequentially digested in two digestion buffers (PBS containing 200 U/ml collagenase I (SCR103, Sigma Aldrich), 0.05 U/ml elastase (E1250, Sigma Aldrich), 5 U/ml neutral protease (LS02111, Worthington), and 0.3 U/ml deoxyribonuclease I (M6101, Promega) for 20 min, followed by DMEM containing 5 mg/ml collagenase type II [C6885, Sigma-Aldrich] and 0.5 mg/ml elastase [LS002292, Worthington Biochemistry]) for 2.5 min at 37 °C). The tissue suspension was filtered with a 40μm cell strainer, then centrifuged at 500 g for 5 min. Cells were resuspended with PBS containing 0.04% BSA. Single-cell suspensions from 6 DVT-adjacent IVCs were pooled to generate the experimental sample (DVT), and suspensions from 6 sham IVCs were pooled to generate the control sample (SHAM). 10,000 cells were loaded using the 10x Genomics Chromium platform with the Single Cell 3' v3.1 Reagent Kit to construct libraries. Libraries were then sequenced on an Illumina NovaSeq X Plus flowcell using a 2×150 bp sequencing reaction, targeting >90,000 reads per cell.