title: G1_biological_replicate_3
design: Ribosome protected footprints were generated through nuclease treatment of cell extract in the presence of cycloheximide. The post-mitochondrial supernatant of the lysate was loaded onto 2 mL of 33% Sucrose as the lower cushion and ultracentrifuged for 2 hours 45 minutes at 35,000 RPM. Following centrifugation, the ribonuclease protection assay was carried out using 250 units of Benzonase for 10 minutes at 25C on the polysomes pellet. Treated RNA was extracted (mirVana miRNA Isolation Kit, Thermo Fisher) and ribosome-protected fragments (aprox. 30 nt) were separated and purified through FlashPAGE electrophoresis.  Library preparations were carried out using TruSeq RNA Library Prep Kit v2 (Illumina), following the manufacturer instructions.