Identification of Alternaria brassicicola genes expressed in planta during pathogenesis of XXXX .

@entity958 @entity281 @entity3789 @entity3494 @entity5189
Alternaria brassicicola is a necrotrophic fungal pathogen that causes black spot disease on cruciferous plants including economically important Brassica species. The purpose of this study was to identify fungal genes expressed during @entity281 of @entity958 . In order to identify candidate genes involved in pathogenicity, we employed suppression subtractive hybridization (SSH) between RNA isolated from A. brassicicola spores incubated in water and on the leaf surface of the @entity958 ecotype Landsberg. Two populations of cDNA were created from total RNA extracted after 24h when approximately 80% of the spores had germinated either on the leaf surface or in water. Following SSH, expression of clones was examined using dot-blot macro-arrays and virtual Northern blots. 47 cDNA clones differentially expressed between Alternaria infected @entity958 leaves and spore germination in water were selected for sequencing. Seventy-seven percent (36) of the cDNAs had significant homology to fungal sequences from databases examined, including available fungal genomes, while 13% (11) had no homology to sequences in the databases. All 36 genes had significant matches with genes of fungal origin, while 11 genes did not have significant hits in the databases examined. Five sequences were expressed on the plant leaf surface but not during spore germination in water according to virtual Northern blots. These five cDNAs were predicted to encode a @entity3789 hydratase, @entity3494 ATPase, @entity5189 dehydrogenase, major Alternaria allergen, and one unknown. RT-PCR was used to examine the expression of these five genes during @entity281 of Brassica oleraceae var. capitata (cabbage), in vitro growth in nutrient rich media, and @entity281 of @entity958 . Four of these genes are expressed in the nutrient rich medium, while the unknown gene P3F2 was only expressed during @entity281 . The results of this study provide the first insight into genes expressed during A. brassicicola @entity281 of Brassica species that may be involved in fungal pathogenesis.
