GSM6597867: high-fat diet, replicate 1, scRNAseq; Mus musculus; RNA-Seq
aortas were cut and digested with an enzyme solution (0.2% type IA collagenase [Sigma, USA] and 200 U/ml DNase [Sigma, USA]) for 30 min at 37°C. The cell suspension was strained through a 40-μm filter (STEMCELL Technologies, China), treated with ACK lysis buffer for 1 minute at room temperature (Sigma, USA), and washed thrice with PBS. The cells were resuspension in PBS with 0.04% bovine serum albumin (Beyotime, China), and their viability was greater than 80%. Qualified aortic cell suspensions (500–1000 living cells per microliter as determined by Count Star) were loaded onto a Single-cell Controller (10x Genomics GemCode Technology, CA) to generate single-cell gel beads in emulsion (GEMs, Single cell 3′ Library, Gel Bead Kit V2 and Chromium Single Cell B Chip Kit [10x Genomics]). Briefly, the single-cell suspensions, gel beads, reagents and partitioning oil were loaded onto each channel. Single-cell RNA was barcoded through reverse transcription in individual GEMs. Barcoded cDNA was pooled, and a general library was prepared using Single Cell 3′ Library and Gel Bead Kit V2. The library was finally sent for sequencing.