Laboratory of Molecular Cytogenetics and Cytometry
Institute of Experimental Botany AS CR

Positional cloning in a huge allopolyploid genome of bread wheat was considered infeasible, mainly due to presence of three homoeologous genomes and large amounts of repetitive sequences. One of strategies enabling to reach this target is based on the use of genomic resources derived from parts of the hexaploid genome. Here we propose construction of two subgenomic BAC libraries specific for short and long arm of chromosome 7D. They will be invaluable resource for creation of physical maps of chromosome 7D and map-based cloning of several agronomically important genes from chromosome 7D of wheat, including genes conferring resistance to green bug, powdery mildew, stem rust, leaf rust, stripe rust, Septoria tritici blotch and Russian wheat aphid. The project will focus on positional cloning of gene Dnx for resistance to Russian wheat aphid (RWA) by creating high-density genetic map for the Dnx region and BAC contig spanning this region. Further, a pooled genomic BAC library will be prepared from a RWA resistant line, which will be used to clone the Dnx gene.

Meadow fescue (Festuca pratensis Huds., 2n=2x=14) is perennial, outbreeding grass species carrying genes of winter hardiness and drought tolerance. On the other hand, Italian (L. multiflorum) and perennial ryegrass (L. perenne) which are widely used as turf and forage grasses for their agronomic characteristics, are sensitive to abiotic stresses. The main goal of the project is the construction of a genomic BAC library of meadow fescue and the use of BAC clones for development of a physical cytogenetic map. This map will be integrated with a genetic map. This will establish conditions for more precise physical mapping of genes for resistance to abiotic stresses.

The large size and polyploid nature of the nuclear genome of bread wheat pose a great challenge for the understanding of complete genome information, genome sequencing and gene discovery. This task could be simplified by fractionating the genome into small and defined parts. Here we propose to create two subgenomic BAC libraries specific for short and long arm of wheat chromosome 3D. Their creation will mark the completion of BAC resources for homeologous group 3, which has been selected for the pilot phase of the wheat genome sequencing project. 3D-specifc BAC resources developed in this project will be used to develop a molecular cytogenetic map and, in collaboration with Prof. J. Dvooák (UC, Davis), a physical contig map of 3D. They will also be used for targeted isolation of low-copy (potentially coding) DNA sequences, discovery of gene-rich regions on 3D, and for high-throughput mapping of expressed sequences to chromosome 3D using wheat GeneChip (collaboration with Prof. T. Close, UC, Davis). The se advances will open avenues for comparative analysis of the wheat genome evolution and the analysis of interaction between the genomes of allopolyploid species. The results obtained and genomic resources generated, will allow the isolation and manipulation of important genes to improve tolerance of wheat to biotic and abiotic stresses and quality of plant products.