GSM9512738: subcutaneous white adipose, total RNA, control, rep3; Mus musculus; RNA-Seq
12 week-old male mice were fed chow diet supplemented with 600 mg/kg doxycycline for 2 weeks and SVF cells were isolated from sWAT as described above. SVF was harvested from sWAT as described above. After centrifugation the cells were resuspended in 2% FBS in PBS containing propidium iodide (PI; 1:400) for 15 min incubation on ice. After washing, the cells were resuspended in 2% FBS in PBS for sorting of viable cells, PI- cells. FACS was conducted with a BD Bioscience FACSAria cytometer at the Flow Cytometry Core Facility at UT Southwestern. Total RNA-seq: Total RNA was extracted from adipocytes from control and ad-SMPD3KO mice using RNAqueous Micro Total RNA Isolation Kit (ThermoFisher AM1931), followed by ribosomal RNA (rRNA) depletion with the NEBNext® rRNA Depletion Kit v2 (Human/Mouse/Rat; NEB, E7400). rRNA-depleted RNA was used for library preparation with the NEBNext® Ultra™ RNA Library Prep Kit for Illumina® (NEB, E7530) following the manufacturer's protocols.