Fig. 1

The commonly used single-transcript style versus the RiboPlotR style for a hypothetical gene. A Example of a commonly used single-transcript plot. RNA-seq and Ribo-seq reads are shown for one mature transcript isoform per plot. For Ribo-seq reads, either the most 5′ site or the P-site position (the first nt within the peptidyl site within the ribosome) is used for plotting. The first nt of the transcript is considered frame 1. Reads that are mapped to each reading frame are shown in red, blue, and green. Thus, the Ribo-seq reads for the annotated CDS can be in one of the three colors. AUG start codons are marked as white half-lines in all three frames. The stop codons are marked as grey lines in all three frames. B The RiboPlotR style for the same gene in A shows all annotated transcript isoform models in parallel with RNA-seq coverage and Ribo-seq P-site reads. Within the gene model, the grey boxes indicate 5′ UTRs, the black boxes indicate the annotated ORFs, and the white pentagonal arrows indicate 3′ UTRs. The isoform being considered is labeled in bold. In addition to the annotated ORF, one upstream ORF (yellow box in the gene models) can be shown in the same plot. For all transcript isoforms of a given gene, the same RNA-seq coverage and Ribo-seq P-sites are used for plotting. For the annotated ORF, the expected CDS range is marked between a black dashed line (translation start site) and a grey dashed line (translation stop site); for the uORF, the CDS range is marked between a green dashed line (translation start site) and an orange dashed line (translation stop site). The Ribo-seq P-sites that map to the expected frame, + 1 frame, and + 2 frame are marked in red, blue, and green, respectively. The Ribo-seq P-sites mapped outside of the expected CDS in either the annotated ORF or uORF are shown in grey. Thus, the majority of the Ribo-seq reads for the annotated CDS will be in red if the data agree with the annotation (here the isoform 1 is being considered). From the data, we can infer isoforms 1 and 2 are both expressed and translated, but isoform 3 is not expressed. Since we selected isoform 1 for plotting here, Ribo-seq P-sites that are unique to isoform 2 and not included in isoform 1 are marked in grey