Fig. 4

Separation of operonic genes from monocistronic genes in the absence of specialised spliced leaders (SLs), illustrated with SLOPPR data from the tunicates Ciona intestinalis (top panels) and Oikopleura dioica (bottom panels). All panels display distributions of distances between operonic or non-operonic genes, and labels provide gene numbers. Left panels: in both organisms, a single SL is added to monocistronic and operonic genes, causing SLOPPR to incorrectly designate monocistronic SL-receiving genes with large intergenic distances as operonic. Middle panels: an optimal distance cut-off for operonic genes is inferred via K-means clustering, and genes at or above the cut-off (red notches at 85 and 414 bp respectively) are re-classified as monocistronic non-operonic (red labels). Right panels: a lower manual cut-off (1 bp and 60 bp respectively; red notches at 2 and 61 bp) further reduces the set of genes retained as operonic. Note the peak of tightly spaced non-operonic genes in the O. dioica panels; these genes are likely operonic genes but no SL evidence was obtained from the RNA-Seq data