GSM6927009: Excluded [SAM24376857]; Mus musculus; RNA-Seq
Each tissue biopsy was transferred to a separate 1.5 mL tube (Eppendorf, Germany) containing 0.25 mL Invitrogen TRIzol Reagent (Thermo Fisher Scientific, MA, USA). Biopsies were incubated in TRIzol for 5 min with intermittent vortexing. 50 µL chloroform (MilliporeSigma, MA, USA) was added to the homogenate, vortexed for 20 sec, and incubated at 20-25°C for 2-3 min. To accelerate phase separation, samples were centrifuged at 10,000xg for 18 min at 4°C. The aqueous (top) phase was removed by aspiration and transferred to a clean 1.5 mL tube (Eppendorf, Germany). A volume of 100% RNAse-free ethanol (MilliporeSigma, MA, USA). equal to the volume of the aqueous layer was added, and the RNA was further isolated using a RNeasy Micro Kit (Qiagen, Germany). Alternatively, an individual tumor biopsy was immersed in RNA later and the RNA further extracted using a RNeasy Mini Kit (Qiagen, Germany).