A preliminary study of changes in the proteome of annual ryegrass (Lolium rigidum Gaud.) seeds during dormancy release is described in this chapter. Highly dormant seeds (i.e. light-insensitive, unable to germinate under light or dark conditions) were prepared by stratification in darkness at 20 C for 2 days. Seeds with a low level of dormancy (i.e. fight-sensitive, able to germinate under fight but not dark conditions) were prepared by stratification in darkness at 20 degrees C for 35 days. 2D gel electrophoresis (2D-PAGE) was performed using excised embryos. Only six proteins were found to differ in abundance between light-insensitive and fight-sensitive embryos. Spots I and 2 were too low to be analysed in light-insensitive embryos but were present in fight-sensitive embryos, representing at least a 20-fold induction. Spots 3 and 4 were three- to fivefold more abundant in fight-insensitive compared with fight-sensitive embryos. Spots 5 and 6 were more abundant in light-sensitive embryo samples, but this difference was less than twofold. The six protein spots of interest were excised from the gels, digested with trypsin and analysed by tandem mass spectrometry (MS/MS). Spots 1 and 2 gave significant matches with glyceraldehyde 3-phosphate dehydrogenases (GAPDH). Spot 4 could not be matched either against the NCBInr Viridiplantae (Green Plants) or a translation of the Lolium gene index from The Institute for Genomic Research (TIGR). De novo sequencing of two peptides was performed, but matching of these sequences to Lolium or total plant protein data-sets with the basic local alignment search tool (BLAST) has been unsuccessful. Data from spots 3, 5 and 6 were not sufficient to allow partern matching identification, or successful de novo sequencing analysis, and remain unidentified.