Characterization of the prehaustorial resistance against leaf rust (Puccinia triticina f. sp. tritici) in Einkorn (Triticum monococcum) by massive analysis of cDNA ends (MACE)

Leaf rust caused by Puccinia triticina f. sp. tritici is the most common rust disease of wheat and causes high yield losses worldwide. Triticum monococcum accessions are valuable sources for improving leaf rust resistance in hexaploid wheat. In extensive screening programs T. monococcum accession Pi272560 has been identified showing prehaustorial resistance against leaf rust. First experiments revealed that the effective defense reaction is associated with an increased activity of peroxidases and pathogenesis related genes. This race non-specific (horizontal) prehaustorial resistance prevents the infection prior to the formation of haustorial mother cells. Hence the goals of our studies are (i) to analyze the biochemical background of this resistance by microscopy and measurement of the H2O2 content in leaves and (ii) to determine the molecular background by genome wide expression studies using the massive analysis of cDNA ends (MACE). A xylenol orange assay revealed higher amounts of H2O2 up to 1.88 µM g-1 in inoculated leaves 12 to 48 hours after inoculation (hai) in the resistant accession. In order to analyze the expression of genes which led to observed defense reactions, MACE from RNA samples which were isolated within the first 24 hai from resistant and susceptible T. monococcum accession has been conducted. Within the time segment from 0-8 hai 120950 tags, between 8-16 hai 95147 tags, and between 16-24 hai 90150 tags could be annotated to the Swissprot database. Using the counts per million determined tags (cpm) up to 8 hai 423, between 8-16 hai 523, and between 16-24 hai 552 tags differentially expressed were identified. These tags have been blasted to the NCBI database and differentially expressed peroxidases (between 9-31), chitinases (2-13), kinases (32-80) and pathogenesis related genes (0-18) were determined. The results show that higher or exclusive expression of peroxidases, chitinases and pathogenesis related genes are involved in prehaustorial resistance. In addition seven genes related to (leaf rust) resistance genes were detected. Based on the analysis of 1136 tags differentially expressed comprising 4358 SNPs, 362 differentially expressed SNP containing genes were mapped in silico using the Genomezipper.