Recall experimental validated TSS. Comparison of 74 experimentally validated TSS described in literature  with TSSAR results. The Manual TSS annotation recovered 40, 15 and 6 TSS with a 0, ±1 and ±2 nt offset, respectively. Here 12 TSS were annotated more than 10 nt away from the experimentally determined position (summarized as missed in the plot). TSSAR was run with a Sensitive and a Specific parameter set (p-value cutoff 0.05 and 0.0001; noise cutoff 1 and 3, respectively). With sensitive parameters 39 TSS (53%) were annotated on the exact same position. Of the remaining TSS 13 and 7 were annotated with ±1 and ±2 nt variance, respectively, whereas 14 TSS (19%) were annotated more than 10 nt away. The specific TSSAR prediction annotated 37, 9 and 6 TSS with 0, ±1 and ±2 nt offset, respectively, relative to the experimentally validated position. In this case 21 TSS (28%) were annotated more than 10 nt away, and therefore annotated as missed. The results of the same analysis including also our naïve benchmark approaches can be found in Additional file 1: Figure S3.