Abstract
Although the human gene annotation has been continuously improved over the past 2 decades, numerous studies demonstrated the existence of a "dark proteome", consisting of proteins that were critical for biological processes but not included in widely used gene catalogs. The Genotype-Tissue Expression project generated more than 15,000 RNA-seq datasets from multiple tissues, which modeled 30 million transcripts in the human genome. To provide a resource of high-confidence novel proteins from the dark proteome, we screened 50,000 mass spectrometry runs from over 900 projects to identify proteins translated from the Genotype-Tissue Expression transcript model with proteomic support. We also integrated 3.8 million common genetic variants from the gnomAD database to improve peptide identification. As a result, we identified 170,529 novel peptides with proteomic evidence, of which 6048 passed the strictest standard we defined and were supported by PepQuery. We provided a user-friendly website (https://ncorf.genes.fun/) for researchers to check the evidence of novel peptides from their studies. The findings will improve our understanding of coding genes and facilitate genomic data interpretation in biomedical research.