GSM8645771: TBLC_GLORI_siVirma_rep1; Mus musculus; RNA-Seq
TRIzol reagent (Invitrogen, 15596026) was added to cell samples to extract RNA. Briefly, 0.5 mL TRIzol reagent was added to wells in a 6-well plate and mixed thoroughly. Next, 0.2 mL chloroform was added (for phase separation), followed by centrifugation at 12,000 g at 4°C for 15 min. The upper aqueous phase was transferred to a new RNase-free tube and an equal volume of isopropanol added. After mixing, RNA was precipitated for 2 h at -20°C, and then centrifuged for 10 min at 12,000 g at 4°C. RNA pellets were washed twice in 75% ethanol and dissolved in RNase-free water. mRNA purification was performed as described; 50 ng mRNA was used as a starting material following a previous GLORI library protocol. Libraries were sequenced on an Illumina NovaSeq 6000 platform (JMDNA (Shanghai) Bio-Medical Technology Co., Ltd.).