Total cellular RNA was extracted using hot acid-phenol extraction and the resulting RNA was chemically fragmented. Samples were barcoded using a 3’ adaptor with a unique sequence corresponding to each sample, and then pooled for downstream processing, as described in Shishkin et al., 2015 (link). rRNA was depleted from the pooled samples using the Illumina Ribo-Zero Gold rRNA Removal Kit for Yeast (MRZY1306). Sequencing was performed at the Functional Genomics Core at the University of Chicago. Detailed protocol for library preparation is available; see Shishkin et al., 2015 (link).
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