GSM6208432: Exosc3_mESC_CTRL_TTseq_Rep1; Mus musculus; OTHER
For RNA extraction, cells were washed 3x with DPBS and lysed directly on the plate with 1mL of TRIzol reagent (Invitrogen, 15596026). Subsequently, RNA was purified using Direct-zol RNA Miniprep Plus Kits (Zymo, R2072) following manufacturer instructions. In brief, lysed cells in TRIzol were mixed 1:1 with 100% ethanol. Mixed lysates were added to the Zymo-spin columns, treated with DNase I, and washed with provided buffers. RNA was eluted with UltraPure DNase/RNase-Free Distilled Water (Invitrogen, 10977015). 250 ng of enriched RNA was used for library construction with the Illumina TruSeq stranded total RNA kit and Illumina CD index plate. After 5 cycles of PCR, samples were removed from the thermal cycler and a test PCR was performed to determine the optimal number of final cycles.