Secreted proteins from plants and phytopathogens play important roles in their interactions and contribute to elaborate mechanisms of attack, defense, and counter-defense, as well as surveillance and signaling. There is therefore considerable interest in developing techniques to characterize "secretomes." Here, we describe the use of the yeast secretion trap (YST) functional screen to isolate and identify secreted proteins that are accumulated and detected in the extracellular matrix of eukaryotes. This method involves fusing cDNAs generated or derived from plants, pathogens, or infected tissue to a yeast (Saccharomyces cerevisiae) invertase (suc2) reporter gene lacking its signal peptide, transforming the resulting fusion library into an invertase-deficient yeast strain, and plating the transformants on a sucrose selection medium. A yeast transformant containing a cDNA that encodes a secreted protein can rescue the mutant and the plasmid DNA can then be sequenced to identify the secreted protein. The YST screen can be a very powerful tool in the study of dynamics of plant host-pathogen interactions.