GSM5493812: Ago2_RS_cko_1_RNA; Mus musculus; RNA-Seq
Cell pellets were homogenized in 1ml of TRIzol reagent (Invitrogen, 15596026) and incubated at room temperature for 5 mins. After incubation, 200µl of chloroform was added followed by vortexing, brief incubation at room temperature, and centrifugation at 12000xg for 15 mins at 4°C. The resultant aqueous phase was transferred to a new tube and mixed with an equal volume of 100% ethanol, then transferred to an RNeasy MinElute spin column (Qiagen, 74104). RNA was extracted on the column according to the manufacturer's instructions. The quality of the eluted RNA was determined by measuring 28S/18S ribosomal ratios and RNA integrity number (RIN) using an Agilent Bioanalyzer. RNA sample integrity was assessed by BioAnalyzer (Agilent Technologies), and samples with RIN >7 were used for paired-end sequence library construction using the KAPA mRNA HyperPrep Kit (Roche # 08098123702).