Ascorbate (vitamin C) is a powerful antioxidant and scavenger of free radicals that protects plants against oxidative damage caused by adverse environmental conditions such as drought, salt stress, and herbicide use. Dehydroascorbate reductase (DHAR; EC 1.8.5.1) and monodehydroascorbate reductase (MDAR; EC 1.6.5.4) are crucial for ascorbate regeneration and the maintenance of a pool of the reduced form. In this study, we report the development of transgenic potato (Solanum tuberosum L.) that overexpresses the Arabidopsis thaliana DHAR gene (AtDHAR1) in the cytosol. The transgenic plants exhibited up to 4.5 times the DHAR activity and up to 2.8 times the level of reduced ascorbate found in the wild-type plants. When subjected to methylviologen treatment, the transgenic plants exhibited enhanced tolerance in terms of less ion leakage, greater chlorophyll contents, less accumulation of hydrogen peroxide, and less severe visual injury symptoms. Moreover, the transgenic plants exhibited faster growth under drought and salt stress. Our results demonstrate that elevating ascorbate contents by overproducing DHAR represents a viable approach for the development of herbicide-tolerant potato.Key Words: ascorbate, potato, vitamin C, dehydroascorbate, methylviologen. IntroductionAdverse environmental conditions such as drought, salt stress, and use of herbicide promote the overproduction of reactive oxygen species (ROS) in plant cells. ROS such as singlet oxygen (O 2 1 ), superoxide radicals (O 2 ⋅), hydrogen peroxide (H 2 O 2 ), and hydroxyl radicals (OH⋅) are believed to be the major factor responsible for rapid cellular damage due to their high reactivity with membrane lipids, proteins, and DNA (Asada and Takahashi 1987, Hung et al. 2005, Mittler 2002). Scavenging and reducing the deleterious effects of ROS is performed by several enzymatic and nonenzymatic systems in plant cells. A network of low-molecularweight antioxidants, such as ascorbate (AsA), glutathione (GSH), and α-tocopherol, and antioxidative enzymes such as superoxide dismutase (SOD), ascorbate peroxidase (APX), and catalase (CAT) are crucial for preventing and controlling the dangerous affects of ROS (Noctor and Foyer 1998). Specifically, enzymes involved in catalyzing redox reactions and in maintaining pools of reduced AsA and GSH in the AsA-GSH pathway, mainly monodehydroascorbate reductase (MDAR; EC 1.6.5.4), dehydroascorbate reductase (DHAR; EC 1.8.5.1), and glutathione reductase, are considered of paramount importance in plant antioxidant defense mechanisms.The antioxidant AsA can directly scavenge free radicals (Halliwell and Gutteridge 2000), and is particularly important as an electron donor for detoxification of H 2 O 2 via ascorbate peroxidase in plant cells (Noctor and Foyer 1998). Moreover, AsA is a major redox buffer in plants , a cofactor of many enzymes (Smirnoff and Wheeler 2000), a regulator of cell division and growth (Kerk and Feldman 1995), and a molecule used in signal transduction in plants (Noctor et al. 2000). AsA is synthesized in t...

The tolerance of the dune grass Leymus mollis (Triticeae; Poaceae) to various biotic and abiotic stresses makes it a very useful genetic resource for wheat breeding. Wide hybridization between L. mollis and wheat allows the introduction of Leymus chromosomes into the wheat genetic background and facilitates the integration of useful traits into wheat. However, the genetic basis controlling the physiological tolerance of L. mollis to multiple environmental stresses remains largely unexplored. Using suppression subtractive hybridization, we identified 112 osmotic-stress-responsive genes from L. mollis and confirmed their differential expression under osmotic stress. These genes were categorized into 13 functional categories, including cell defense and stress response, transcriptional regulation, signal transduction, biosynthesis of compatible solutes and cell wall metabolism. Representative genes were validated by northern blot and RT-PCR analyses of expression patterns in response to osmotic stress and abscisic acid treatment. The genes identified here represent a useful source of expressed sequence tags (ESTs) for the analysis and identification of Leymus chromosomes introduced into wheat. Furthermore, being highly conserved, genetically associated with osmotic stress tolerance and transferable to wheat, these ESTs provide significant tools for the development of EST-derived molecular markers for introgression of osmotic stress tolerance genes into wheat.

Potato (Solanum tuberosum L.) is the world's number one non-cereal food crop and ranks fourth among most important crops grown worldwide in terms of acreage, yield and value. In order to maintain greater protection against environmental stresses, we developed transgenic potato overexpressing Arabidopsis thaliana glutathione reductase gene (AtGR1). The transgenic potato maintained up to 6.5 folds higher GR activity, 5.8 folds glutathione (GSH) contents and up to 2.2 folds higher glutathione S-transferase activity compared to non transformed plants (NT). Interestingly, while the transgenic plants exhibited decreased dehydroascorbate reductase (DHAR) activity, the relative reduced ascorbate (AsA) contents were higher while the relative dehydroascorbate (DHA) were lower compared to NT which provide a support to the hypothesis that an active glutathione-independent pathway for DHA reduction might exists in vivo. The transgenic plants maintained an enhanced tolerance to methylviologen, and cadmium. When subjected to drought stress, the transgenic plants exhibited faster recovery with less visual injury compared to NT. These results suggest that manipulation of glutathione levels provides reliable strategy for the development of industrial transgenic potato plants with enhanced tolerance to multiple environmental stresses.

Leymus mollis (Triticeae; Poaceae) is a useful genetic resource for wheat (Triticum aestivum L.) breeding via wide hybridization to introduce its chromosomes and integrate its useful traits into wheat. Leymus mollis is highly tolerant to abiotic stresses such as drought and salinity and resistant to various diseases, but the genetic mechanisms controlling its physiological tolerance remain largely unexplored. We identified and cloned an allene oxide cyclase (AOC) gene from L. mollis that was strongly expressed under salt stress. AOC is involved in biosynthesis of jasmonic acid, an important signaling compound that mediates a wide range of adaptive responses. LmAOC cDNA consisted of 717 bp, coding for a protein with 238 amino acids that was highly similar to AOCs from barley (Hordeum vulgare) and other monocots. Subcellular localization using Nicotiana benthamiana confirmed it as a chloroplast-localized protein. LmAOC was found to be a multiple-copy gene, and that some copies were conserved and efficiently expressed in wheat–Leymus chromosome addition lines. LmAOC expression was upregulated under drought, heat, cold and wounding stresses, and by jasmonic acid and abscisic acid. Our results suggest that LmAOC plays an important role in L. mollis adaptation to abiotic stresses and it could be useful for wheat improvement.

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