Poly a polymerase kit

SNU-Seq libraries were generated by following the TT-Seq protocol (Schwalb et al., 2016) , but omitting the sonication step. Before library preparation, nascent RNAs were polyadenylated using the NEB Poly(A)-Polymerase kit following the kit instructions. The reaction with 150 ng thiolabeled RNA was left for 45 minutes at 37 ˚C before isopropanol precipitation. The pellet was resuspended in 11 L RNase-free water. Qualities and amounts were checked on the Qbit fluorometer and the Agilent Bioanalyzer (RNA Pico Chip). 6x10 6 Hep3B cells were treated with 10 ng/ml IFN for 0, 0.5, 2 or 24 hrs. During the final 12 min of IFN treatment, 500 M 4sU was additionally added. Following treatments, cells were washed with cold PBS before lysing in Qiazol on ice. Samples were spiked with Saccharomyces cerevisiae RNA (0.01%). 300
g RNA was biotinylated.
Libraries were prepared following the Quant-Seq Lexogen 3' mRNA kit instructions with 13 PCR cycles. Library qualities were checked using the Agilent Bioanalyzer (DNA High Sensitivity Chip). Following the manufacturer's instructions, the chips prepared by the Ion Chef were then sequenced on an Ion Proton Sequencing platform.