GSM4932129: HR_Cntrl_NAT10_rep1; Homo sapiens; OTHER
Input RNA was obtained from 130 μL aliquots of lysate (without RNase I digestion) using TRIzol Reagent. Total RNA was treated with Turbo DNase I (ThermoFisher, 30 min at 37°C). RNA Spike-In mix (ThermoFisher Scientific, Cat. #: 4456740) was added to 5 μg of DNase I-treated total RNA according to the manufacturer's instructions, and ribosomal RNA was depleted using the human/mouse/rat Ribo-Zero Gold rRNA removal kit. Next, total RNA samples were fragmented in alkaline fragmentation buffer (1 mM EDTA, 50 mM Na2CO3, 50 mM NaHCO3, [pH 9.2] for 40 min at 95°C, as described previously. Fragments between 26 and 34 nucleotides in size were gel purified from input and footprinted RNA samples, and used for library preparation as described previously. Sequencing libraries were constructed with the Illumina TruSeq Stranded Total RNA Library Prep Kit (RS-122-2201).