title: GSM4809427: V6.5_ES_epiLC_96hr_shBcat_rep1; Mus musculus; RNA-Seq
library_construction_protocol: RNA was isolated by first homogenizing cells in 1ml TRIzol reagent (Invitrogen 15596026) through triturating the cell lysate through a 27-gauge needle a minimum of 7 times. Homogenized samples were mixed with 200µ l chloroform and vigorously shaken by hand for 30sec. Samples were then incubated at RT for 10min before centrifuging at 12,000g for 10min at RT. The aqueous phase containing RNA was transferred to a new tube, mixed with 100µ l chloroform, and subjected to a second round of shaking, incubation and centrifuging.  The aqueous phase was transferred to a new tube containing 1.5µ l 20mg/ml glycogen, and mixed with 450µ l isopropanol to pre- cipitate RNA. Samples were gently mixed and incubated at RT for 10min before centrifuging at 4 °C for 15min. The supernatant was discarded and the pellet was washed by 1ml 75% ethanol. After the wash the RNA pellet was dried at RT for 20min and resuspended in RNase-free water. Yield and purity of RNA was determined by spectrophotometry with Nano drop (Thermo Scien- tific). The RNA was treated with DNase (Promega M6101) RNA-seq and ChIP-seq libraries were prepared using the TruSeq RNA kit for sequencing,