GSE94460
GSE GEO Potentially relatedRibosome profiling of HEK293 cells.
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Summary
The development of the ribosome profiling (ribo-seq) technique has enabled the measurement of translation at a genome-wide level. There are several variants of the ribosome profiling technique that use different translation inhibitors. The regular ribo-seq utilizes Cycloheximide (CHX), a translation elongation inhibitor to freeze all translating ribosomes. In contrast to CHX, the translation inhibitor lactimidomycin (LTM) and harringtonine (Harr) have a much stronger effect on initiating ribosomes. The use of these inhibitors allows for the global mapping of translating initiating sites (TISs) when they are coupled with ribosome profiling (TI-seq). We have developed a computational tool to detect and/or quantitatively compare translation initiation from TI-seq data, and predict novel ORFs from regular CHX-based ribo-seq data. Two replicates of CHX-based ribo-seq experiments and one Harr based ribo-seq were performed in HEK293 cells for confirming the ORFs predicted using public available data.
Overall Design
Ribosome protected fragments were extracted from HEK293 cells treated with cycloheximide or harringtonine plus cycloheximide.
Analysis (4 steps)
View Data Processing- Library strategy: Ribosome profiling
- Fastq reads were trimmed and mapped to human rRNA sequences using bowtie allowing 2 mismatches.
- Unmapped reads were mapped to human genome GRCh38 with Ensembl gene annotation release 83 using STAR with parameters â--outSAMattributes All --outFilterMismatchNmax 2 --alignEndsType EndToEnd --outFilterIntronMotifs RemoveNoncanonicalUnannotated --alignIntronMax 20000 --outMultimapperOrder Random --outSAMmultNmax 1â
- Reads with mapQ=0 or NH>5 were filtered out and only primary loci are counted.
Supplementary Files (1)
GEO Samples (3)
Dataset Citations (1)
Potentially Related Publications (1)
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