Total RNA was extracted by a hot-phenol protocol and then mRNA was isolated with Poly(A)Purist MAG kit (Ambion). RPF were isolated from the monosomal pic (80S) coming from digestion of polysomes with RNaseI. Next, a denaturing urea gel was used to isolate the RNA fragments corresponding to ribosome footprints (Ingolia, 2009) and (Nedialkova, 2015). Libraries were prepared using the protocol described in (Ingolia, 2009) and (Nedialkova and Leidel, 2015). For RNA-seq libraries, RNA integrity was assessed using a denaturing formaldehyde gel and mRNA isolated using the Poly(A) purist MAG kit (Ambion). For RPF libraries, ribosome footprints were size-selected and purified from a TBE-urea denaturing gel. Libraries were sequenced with Illumina HiScanSQ.
GEO Accession
GSM2958764