Determining Alternative Protein Isoform Expression Using RNA Sequencing and Mass Spectrometry

STAR Protoc. 2020 Oct 21;1(3):100138. doi: 10.1016/j.xpro.2020.100138. eCollection 2020 Dec 18.


Alternative splicing greatly expands the coding capacity of the human genome, but how many alternative transcripts are translated as proteins or carry functional importance remains unknown and awaits experimental verification. Here, we describe a protocol that combines transcriptomics (RNA-seq) and proteomics (mass spectrometry [MS]) analyses to identify alternative isoforms in proteomes. This workflow is applicable to custom-generated RNA-seq and MS data from matching samples, as well as the reanalysis of existing transcriptomics and proteomics datasets in public repositories. For complete details on the use and execution of this protocol, please refer to Lau et al. (2019).

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Alternative Splicing / genetics
  • Base Sequence / genetics
  • Gene Expression / genetics
  • Gene Expression Profiling / methods*
  • Humans
  • Mass Spectrometry / methods*
  • Protein Isoforms / analysis*
  • Protein Isoforms / genetics
  • Protein Isoforms / metabolism
  • Proteome / genetics
  • Proteomics / methods
  • RNA-Seq / methods
  • Sequence Analysis, RNA / methods
  • Tandem Mass Spectrometry / methods
  • Transcriptome / genetics


  • Protein Isoforms
  • Proteome