GSM9036725: 4741_Treated_1_RNAseq; Saccharomyces cerevisiae; RNA-Seq
For RIBO-seq, lysates were prepared according to the procedure and were digested with RNase I for 45 min at room temperature as previously described in McGlincy and Ingolia, 2017. Digestion was stopped with SuperaseIN and monosomes purified by size exclusion chromatography on MicroSpin S-400 HR columns (GE Healthcare) as described in Aeschimann et al., 2015. Size selection of footprints with length 25-33 nt was performed by electrophoresis on 15% TBE-urea gels. For RNA-seq, total RNA was purified from an aliquot of cell lysate and rRNA depleted using NEB NEBNext rRNA depletion kit v2 following manufacturer instructions. mRNA fragmentation was conducted for 20 min at 94°C to generate RNA fragments of sizes similar to those of the ribosome footprints. Illumina ready RIBO-seq libraries were prepared using a SMARTer smRNA-seq kit (TakaraBio).