Genome Biology201920:112 https://doi.org/10.1186/s13059-019-1710-7 © The Author(s). 2019 Published: 3 June 2019 Abstract Genetic, transcriptional, and post-transcriptional variations shape the transcriptome of individual cells, rendering establishing an exhaustive set of reference RNAs a complicated matter. Current reference transcriptomes, which are based on carefully curated transcripts, are lagging behind the extensive RNA variation revealed by massively parallel sequencing.