Fusarium graminearum exploits ethylene signalling to colonize dicotyledonous and monocotyledonous plants.

Ethylene signalling affects the resistance of dicotyledonous plant species to diverse pathogens but almost nothing is known about the role of this pathway in monocotyledonous crop species. Fusarium graminearum causes Fusarium head blight (FHB) of cereals, contaminating grain with mycotoxins such as deoxynivalenol (DON). Very little is known about the mechanisms of resistance/susceptibility to this disease. Genetic and chemical genetic studies were used to examine the influence of ethylene (ET) signalling and perception on infection of dicotyledonous (Arabidopsis) and monocotyledonous (wheat and barley) species by F. graminearum. Arabidopsis mutants with reduced ET signalling or perception were more resistant to F. graminearum than wild-type, while mutants with enhanced ET production were more susceptible. These findings were confirmed by chemical genetic studies of Arabidopsis, wheat and barley. Attenuation of expression of EIN2 in wheat, a gene encoding a core component of ethylene signalling, reduced both disease symptoms and DON contamination of grain. Fusarium graminearum appears to exploit ethylene signalling in both monocotyledonous and dicotyledonous species. This demonstration of translation from model to crop species provides a foundation for improving resistance of cereal crops to FHB through identification of allelic variation for components of the ethylene-signalling pathway.

A comparison of transgenic barley lines produced by particle bombardment and Agrobacterium-mediated techniques.

Two barley transformation systems, Agrobacterium-mediated and particle bombardment, were compared in terms of transformation efficiency, transgene copy number, expression, inheritance and physical structure of the transgenic loci using fluorescence in situ hybridisation (FISH). The efficiency of Agrobacterium-mediated transformation was double that obtained with particle bombardment. While 100% of the Agrobacterium-derived lines integrated between one and three copies of the transgene, 60% of the transgenic lines derived by particle bombardment integrated more than eight copies of the transgene. In most of the Agrobacterium-derived lines, the integrated T-DNA was stable and inherited as a simple Mendelian trait. Transgene silencing was frequently observed in the T1 populations of the bombardment-derived lines. The FISH technique was able to reveal additional details of the transgene integration site. For the efficient production of transgenic barley plants, with stable transgene expression and reduced silencing, the Agrobacterium-mediated method appears to offer significant advantages over particle bombardment.

An efficient method for the physical mapping of transgenes in barley using in situ hybridization.

The genetic transformation of crops by particle bombardment and Agrobacterium tumefaciens systems have the potential to complement conventional plant breeding programmes. However, before deployment, transgenic plants need to be characterized in detail, and physical mapping is an integral part of this process. Therefore, it is important to have a highly efficient method for transgene detection by fluorescence in situ hybridization (FISH). This study describes a new approach, which provides efficient control of probe length and labelling, both of which play an important role in in situ hybridization of transgenes. The approach is based on reducing the size of the plasmid prior to labelling by nick translation, rather than using the whole or linearized plasmid, or varying the amounts of DNaseI in the nick translation mixture. This provided much more efficient labelling of the probe, which yielded optimal hybridization. minimal fluorescent background, and accurate physical location of the transgene.