GSM7634510: 72 hr 3; Mus musculus; RNA-Seq
Cells were centrifuged and cell pellets resuspended in 1 mL Red Blood Cell Lysing Buffer Hybri-MaxTM (Product No R7767, Sigma-Aldrich, St. Louis, MO 63103) and incubated for 5 min at room temperature, then washed with HBSS supplemented with 2% FBS. Viable cells were enumerated using a hemocytometer with trypan blue dye exclusion. Total RNA was extracted from cells by phenol-chloroform extraction method using TRIzol RNA Isolation Reagent (Product No 15596026, ThermoFisher Scientific). Purified RNA was resuspended in 15 uL RNAsecureTM RNase Inactivation Reagent (Thermo Fisher Scientific) and stored at -80 °C. RNA integrity numbers (RINs) were confirmed to be ≥ 8.8 using a 2100 Bioanalyzer Instrument (Agilent, Santa Clara, CA). cDNA libraries were prepared using mouse TruSeq® Stranded Total RNA Library Prep kit (illumina, San Diego, CA) and quantified using a KAPA Library Quantification kit (Roche, Pleasanton, CA).