Improving nitrogen use efficiency in barley (Hordeum vulgare L.) through the cisgenic approach

Barley is one of the major crops cultivated worldwide and constitutes an important basis for animal feed. However, the production is facing a number of challenges that will be accentuated in the years to come, in particular restrictions on the use of nitrogen (N) fertilizer. In order to improve the N use efficiency in barley, we are developing a new generation of genetically modified plants based on the concept of cisgenesis. In this approach, plants are transformed only with their own genetic material. The genes encoding the cytosolic isoform of the glutamine synthetase (GS1) and the tonoplast intrinsic protein TIP2, potentially involve in N management and plant growth, have been selected to be transformed into the barley cultivar Golden Promise. The genomic clones comprising 1-2kb of the promoter, the gene itself and 0.5-1kb of the 3’untranslated region have been isolated and cloned into the pGreenII binary vector. The genes have been inserted into barley by Agrobacterium-mediated transformation using the hygromycin phosphotransferase gene for selection of transformed lines on hygromycin. In this system, the resistance gene is placed on the helper plasmid pSoup allowing for separate introductions of the gene of interest and the resistance gene for selection, respectively.The transgenic lines (T0), currently growing in greenhouse will be self pollinated and the molecular, physiological and agronomic characterization of subsequent generations will be undertaken