Technological Advancement Development of a Fusarium graminearum AVymetrix GeneChip for proWling fungal gene expression in vitro and in planta

Recently the genome sequences of several Wlamentous fungi have become available, providing the opportunity for large-scale functional analysis including genome-wide expression analysis. We report the design and validation of the Wrst AVymetrix GeneChip microarray based on the entire genome of a Wlamentous fungus, the ascomycetous plant pathogen Fusarium graminearum. To maximize the likelihood of representing all putative genes (»14,000) on the array, two distinct sets of automatically predicted gene calls were used and integrated into the online F. graminearum Genome DataBase. From these gene sets, a subset of calls was manually annotated and a nonredundant extract of all calls together with additional EST sequences and controls were submitted for GeneChip design. Experiments were conducted to test the performance of the F. graminearum GeneChip. Hybridization experiments using genomic DNA demonstrated the usefulness of the array for experimentation with F. graminearum and at least four additional pathogenic Fusarium species. DiVerential transcript accumulation was detected using the F. graminearum GeneChip with treatments derived from the fungus grown in culture under three nutritional regimes and in comparison with fungal growth in infected barley. The ability to detect fungal genes in planta is surprisingly sensitive even without eVorts to enrich for fungal transcripts. The Plant Expression Database (PLEXdb, http:// www.plexdb.org) will be used as a public repository for raw and normalized expression data from the F. graminearum GeneChip. The F. graminearum GeneChip will help to accelerate exploration of the pathogen–host pathways that may involve interactions between pathogenicity genes in the fungus and disease response in the plant.