Application of molecular biology tools to wild relatives of wheat

Knowledge of how to alleviate drought related yield loss is still limited due to the complexity of both the stress condition and plant responses. Wild emmer wheat (Triticum turgidum ssp. dicoccoides) is a potential source of important drought-resistance genes for its cultivated relatives. This talk aims to understand changes in the gene and protein expression of wild wheat upon exposure to different level of drought stress. Comparative analyses were also perfrmed to see major playes in wild emmer. Specifaclly, we will discuss TMPIT, an integral membrane protein and DRE binding proteins and their putative microRNAs. Analysis of the AP2/ERF DNA-binding domain of TdicDRF1 as a GST-fusion protein and its binding to DRE by electrophoretic mobility shift assay (EMSA) indicate functional differences between wheat DREBs and those characterized in Arabidopsis thaliana. DREB expression increased in drought-stressed roots, correlating with the RT-PCR results, but not in leaf, showing that tissue-specific regulation occurs at the protein level. Hence, the DREB-DRE interaction undergoes subtle multi-level regulation. As for the TMPIT, the protein was expressed and analysed in Escherichia coli and Saccharomyces cerevisiae. Cellular localisation of the protein in the cell was also investigated using an eGFP-tagged form of the protein in S. cerevisiae. Results obtained by confocal laser microscopy indicated that the TdicTMPIT1 tagged with GFP was localised in a membraneous compartment. It is concluded that TdicTMPIT1 is a membrane protein associated with the drought stress response in wild emmer wheat, and so it may be useful for the improvement of modern wheat genotypes. We will also discuss other molecular approaches to have improved understading of drought stree in wild wheats.