GSM4160132: Empty vector control rep 1 [Ribo-seq]; Homo sapiens; RNA-Seq
Cycloheximide-treated cells were lysed and digested with nuclease to generate ribosome footprints (RBFs). Ribosomes and ribosome footprints were isolated by size exclusion with a MicroSpin S-400 column. The cycloheximide-treated cells from the same pool were also used to extract total RNA for RNA-seq libraries. Ribosome profiling libraries and RNA-seq libraries (used for normalizing ribosome footprint (RBF) data) were generated using the TruSeq Ribo Profile (Mammalian) Kit (illumina). The rRNA was depleted from the ribosome footprints and the total RNA using Ribo-Zero rRNA Removal Kit (illumina). The ribosome footprints were size selected using Urea-PAGE before library preparation. The total RNA was fragmented with heat. The size selected RBFs and fragmented total RNA were ligated to a 3′ adaptor, and reverse transcribed. cDNA was purified by Urea-PAGE and circularized. The cDNA library was amplified and barcoded with Illumina-compatible primers.
GEO Accession
GSM4160132