Fig. 5.

Simulation results. a Short-read simulations—proportion of transcript length left uncovered as longer fragments are simulated in a UMI-based library preparation scenario. Short fragments (100–200 bp) leave most of the transcript uncovered by the reads (> 0.75 proportion), while the simulation of longer (> 300 bp) fragments affects transcripts differently depending on their length, hence the growing distributions in the boxplot. b Short-read simulations—multi-isoform genes (MIGs) detected in each 3′ and 5′ end as well as in the Smart-seq simulation are classified in four intervals, according to their individual percentage of resolution. Results shown for neural stem cells (NSCs) only. Intervals gather MIGs for which 0–25, 25–50, 50–75 and 75–100% of their isoforms are resolved. The 3′ end and 5′ end labels only refer to unique molecular identifier (UMI) simulations. Note that Smart-seq data have been plotted twice, in both the 3′ end and 5′ end bar-graph rows, for completeness and to ease visual comparison. c Long-read simulations—the number of genes for multi-isoform genes detected as sequencing depth per cell is progressively lost. The dashed line indicates the number of multi-isoform genes present in the original neural cell transcriptome. A decrease in depth per cell decreases the number of genes for which more than one isoform can be observed. d Long-read simulations—the number of isoform switches detected between neural stem cells and oligodendrocytes in a similar scenario, assuming half of the cells belong to each cell type (i.e. two cells equate to one oligodendrocyte and one NSC). A decrease in sequencing depth per cell not only prevents detection of isoform ratio expression changes (which constitute the majority differences in isoform expression), but also reduces the number of isoform switches that can be observed. The dashed line indicates the number of NSCs versus oligodendrocyte isoform switches detected in the original transcript expression data