Investigating salinity tolerance in alfalfa (Medicago sativa L.): modern synchrotron and molecular approaches

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My long term research goal is to develop alfalfa (Medicago sativa L.) cultivars with tolerance to abiotic and biotic stresses. My short-term (5-year) objectives for this proposal are to investigate how alfalfa tolerates soil salinity at a whole plant level. I assume spatial and temporal distributions of elemental ions and organic solutes in the leaves, stems, and roots of alfalfa will vary between salt tolerant and susceptible cultivars under a salt stress. A salinity tolerant alfalfa must have a unique osmotic regulation through organic molecules or K+ and Ca2+ ions to reduce ion toxicity in the leaves. I also investigate if links exist among gene expression and biochemical changes and stress hormone's concentration in the leaves. *** To test my hypothesis, two alfalfa cultivars with contrasting tolerance to salinity will be exposed to different salt solutions and plants will be sampled periodically following salt treatments. Synchrotron radiation (SR) based phase contrast X-ray imaging, mid-infrared, soft X-ray, and X-ray Fluorescence mapping at the Canadian Light Source (Saskatoon, Canada) will be applied to quantify and localize elemental ions, organic solutes and detect their spatial and temporal distributions in alfalfa tissues. Synchrotron radiation-Fourier transform infrared (SR-FTIR) spectroscopy will be used to map changes in the ratios of carbohydrates, and α-helix to β-sheet protein secondary structures. A transcriptomic analysis of gene expression in alfalfa leaves in response to salt stress will be performed, and based on the RNA-seq analysis, and available genomic information of the model legume, Medicago truncatula, identify candidate genes to salinity tolerance in alfalfa and validate the expression of selected genes.**** Alfalfa is the most widely planted forage legume in the world. Soil salinity is a major abiotic factor that limits alfalfa production in many regions of the world. Knowledge of the biochemical composition of alfalfa tissues under salt stress is critical for the study of stress tolerance physiology, but there is a limited information available on quantification in whole plant level and spatial localization of those compounds at the tissue, cellular and subcellular levels. A comprehensive understanding of salinity tolerance in alfalfa is needed for development of novel cultivars.**** The proposed project has a goal to train Highly Qualified Personnel (HQP) including one M.Sc. student in forage stress physiology and synchrotron application, and one Ph.D. student in alfalfa genomics, two undergraduate thesis students in experimental design, statistics, plant physiology, and basic bio-technology skills. The findings will advance our understanding of the physiology and genetics of salt tolerance in alfalfa. The identification of genes will be useful for breeding new alfalfa varieties with high salinity tolerance.******