Accurate exon–intron structures are essential prerequisites in genomics, proteomics and for many protein family and single gene studies. We originally developed Scipio and the corresponding web service WebScipio for the reconstruction of gene structures based on protein sequences and available genome assemblies. WebScipio also allows predicting mutually exclusive spliced exons and tandemly arrayed gene duplicates. The obtained gene structures are illustrated in graphical schemes and can be analysed down to the nucleotide level. The set of eukaryotic genomes available at the WebScipio server is updated on a daily basis. The current version of the web server provides access to ∼3400 genome assembly files of >1100 sequenced eukaryotic species. Here, we have also extended the functionality by adding a module with which expressed sequence tag (EST) and cDNA data can be mapped to the reconstructed gene structure for the identification of all types of alternative splice variants. WebScipio has a user-friendly web interface, and we believe that the improved web server will provide better service to biologists interested in the gene structure corresponding to their protein of interest, including all types of alternative splice forms and tandem gene duplicates. WebScipio is freely available athttp://www.webscipio.org.