Conservation assessment of human splice site annotation based on a 470-genome alignment.

Despite many improvements over the years, the annotation of the human genome remains imperfect, and different annotations of the human reference genome sometimes contradict one another. The use of evolutionarily conserved sequences provides a strategy for selecting a high-confidence subset of the annotation that is more likely to be related to biological functions, and the rapidly growing number of genomes from other species increases its power. Using the latest whole genome alignment, we found that splice sites from protein-coding genes in the high-quality MANE annotation are consistently conserved across more than 400 species. We also studied splice sites from the RefSeq, GENCODE, and CHESS databases that are not present in MANE. We trained a logistic regression classifier to distinguish between the conservation exhibited by sites from MANE versus sites chosen randomly from neutrally evolving sequence. We found that splice sites classified by our model as conserved have lower SNP rates and better transcriptomic support. We then computed a subset of transcripts only using either "conserved" splice sites or ones from MANE. This subset is enriched in high-confidence transcripts of the major gene catalogs that appear to be under purifying selection and are more likely to be correct and functionally relevant.