Project Number: 5442-21000-033-10
Project Type: Reimbursable

Start Date: Sep 01, 2008
End Date: Aug 31, 2012

Objective:
1. Validate candidate genes for Tsn1 using comparative sequence analysis of EMS-induced Tsn1-disrupted mutants and virus-induced gene silencing (VIGS). The isolation of Tsn1 will provide a necessary tool to gain understanding of toxin recognition by the host. 2. Determine transcriptional structure and expression profiles of Tsn1 using reverse transcription- and relative quantitative-PCR. Molecular characterization of transcriptional expression will provide the fundamental basis for understanding function. 3. Determine levels of allelic diversity at the Tsn1 locus and identify protein domains/amino acids essential for toxin recognition or binding by comparative sequence analysis of Tsn1-disrupted mutants and a large collection of wild-type alleles from diverse genotypes. This will provide clues to critical functional domains and shed light on allelic diversity and evolutionary events leading to toxin sensitivity. 4. Develop a high-resolution map of the Snn7 locus on chromosome arm 5DS in diploid wheat and determine the feasibility of map-based cloning. There is strong evidence indicating that cell death caused by SnTox7 utilizes a different pathway than SnToxA. Therefore, the isolation of Snn7 and characterization of the Snn7-SnTox7 interaction will provide information on different host mechanisms exploited by S. nodorum.

Approach:
The Tsn1 candidate gene region will be defined by positional cloning in a segregating population and haplotype analysis of diverse accessions of wheat relatives. Functional aspects of Tsn1 will be characterized by reverse transcription PCR and quantitative PCR to determine levels of gene expression. Critical domains essential to function will be identified by comparative sequence analysis of a large number of natural and mutated alleles. The fine mapping of Snn5 will be conducted using a large segregating Aegilops tauschii population and the development of markers tightly linked to the gene.