RNA was extracted from yeast cells using the hot phenol procedure. The amount of RNA was quantified in a Nanodrop 2000 Spectrophotometer (Thermo Scientific) and RNA integrity was checked by agarose gel electrophoresis. 5PSeq methods was performed as previously described (PMID: 26820793). 6µg of total RNA was used as input. In brief a RNA oligo (rP5_RND) containing an Illumina adaptor and unique molecular identifiers (UMI) was ligated to the intermediates of mRNA co-translation degradation (5'P). Ribosomal RNA was depleted using Ribo-Zero Magnetic Gold Kit (Illumina).Libraries were PCR amplified (14 cycles). Ampure beads size selected libraries with an average length of 451 nt were sent for sequencing (llumina NextSeq 500 instrument).
GEO Accession
GSM3346538