title: GSM6032712: Exosc3_EpiLC_CTRL_bulkRNA_Rep2; Mus musculus; RNA-Seq
library_construction_protocol: 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). For DNA extraction, PureLink™ Genomic DNA Mini Kit (Invitrogen, K1820-00) was used followed manufacturer instructions. In brief, cells were washed 3x with DPBS and lysed directly on the plates using supplied Lysis/Binding buffer. Lysates were subsequently applied to the column and washed with buffers provided. DNA was eluted in PureLink Genomic Elution Buffer provided in the kit. For complementary DNA synthesis, 100-200 µg of DNase I treated RNA was reverse transcribed using High-Capacity cDNA Reverse Transcription Kit (Applied Biosystems, 4368814). Random hexamers were used to capture total RNA. For metabolic RNA labeling, cDNA synthesis was carried out while RNA was bound to beads using SuperScript VILO cDNA synthesis kit (Invitrogen, 11754-050).