GSM6042622: DF2.ADAR_DF1.IP.flag_Rep2; Homo sapiens; RIP-Seq
Cell lysates (25mM Tris-Cl pH7.4; 150mM NaCl; 2.5mM EDTA, 0.5mM DTT; 0.5% NP-50) were used for RIP using Monoclonal anti-FLAG M2 antibody. Total RNA was extracted from a fraction of the cell lysate (Input samples) or eluted from antibody coupled beads using 1mL of TRIZOL reagent (Thermo) and the RNA was extracts according to the manufacturer's protocol. RNA was resuspended in Nuclease-Free water. Total RNA was extracted using TRIzol reagent. RNA was then subjected to DNaseI treatment to remove any contaminating DNA. For each sample 150ng of total RNA was used for library preparation. Library construction was performed using the NEBNEXT Ultra II Directional RNA-Seq kit and the NEBNext rRNA Depletion Kit.