Description

This track displays ribosome profiles of THP-1 cell lines after treatment with puromycin for the identification of translation initiation sites and treatment with cycloheximide as control published by Jochen Hampe′s lab (Fritsch et al. 2012).

Display conventions

This track contains ribosome profiles from puromycin treated and control samples. The height of the histogram at each location corresponds to the number of reads that match that genomic position.

Methods

THP-1 cells were grown under standard conditions. Cells were treated with Puromycin and incubated to release elongating ribosomes. Translation-initiating ribosomes were arrested by subsequent treatment with cycloheximide. Library preparation was performed as previously described by Ingolia et al. 2009. Footprint fragments were sequenced using the Illumina HiSeq 2000 instrument. Adapter sequences were removed using the short-read alignment software Novoalign V2.07.13 (Novocraft, Selangor, Malaysia) and rRNA contaminations were eliminated by mapping the reads against RefSeq sequences NR_023371, NR_003287, NR_003286, NR_003285 and NR_023363. The remaining reads were aligned to the human reference genome (UCSC GRCh37/hg19 release). The position of the footprint in the ribosome was determined by adding 12 nt to the 5′end of the read. When the first nucleotide of the read failed to align to the reference sequence, 13 nt were added.

Credits

These data were generated and analyzed by the laboratory of Prof. Dr. J. Hampe at the University Hospital Schleswig-Holstein, Kiel, Germany as part of the Virtual Liver Network (contact: Jochen Hampe). This track was constructed by Alexander Hermann at University Hospital Schleswig-Holstein, Kiel.

References

Fritsch, C., Herrmann, A., Nothnagel, M., Szafranski, K., Huse, K., Schumann, F., Schreiber, S., et al. (2012). Genome-wide search for novel human uORFs and N-terminal protein extensions using ribosomal footprinting. Genome Res, 22(11), 2208–2218.

Ingolia, N. T., Ghaemmaghami, S., Newman, J. R. S., & Weissman, J. S. (2009). Genome-wide analysis in vivo of translation with nucleotide resolution using ribosome profiling. Science, 324(5924), 218–223.