GSM9004990: Normoxic mice; Mus musculus; RNA-Seq
RNA was extracted from cells using a Trizol-Chlorofom based method. 1mL of Trizol was added to the cell samples and mixed by inversion then incubated on ice for 5 minutes. 0.2mL of chloroform was added, mixed vigorously and vortexed before incubating at room temp for 3 minutes. Samples were then centrifuged at 11600 x g for 15 mins at 4oC. After which the aqueous phase was transferred to a microcentrifuge tube containing 0.5mL of isopropanol. The sample was pipetted up and down gently 20 times to precipitate the RNA. Samples were incubated at room temp for 10 minutes before being centrifuged at 11600 x g for 10 mins at 4oC. Supernatant was discarded and RNA pellet was washed using 1mL 75% ice-cold ethanol, mixed by inversion and centrifuged at 10000 x g for 5 mins at 4oC. Supernatant was discarded and the RNA pellet was air dried at room temp for 30 minutes. RNA pellet was resuspended in 50uL of nuclease-free water and incubated in a water bath at 60oC for 10 minutes. RNA sequencing libraries were prepared using the NEBNext Ultra II RNA Library Prep Kit for Illumina using the manufacturer's instructions (NEB, Ipswich, MA, USA).