Browsing by Author "Tatlı, Özge"

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    Targeted metabolomic analysis of central carbon metabolism on model plant brachypodium distachyon to elucidate physiological response to drought stress
    (Bahçeşehir Üniversitesi Fen Bilimleri Enstitüsü, 2015-01) Tatlı, Özge; Özdemir, Soğutmaz
    Brachypodium distachyon which is native to the Mediterranean and Middle East Region, is a member of grass family (Poaceae) that also includes a number of cereals, providing the bulk of energy needed for human diet like wheat, barley, rye and oat. B. distachyon, has all desirable features to be a model organism, such as having a compact genome (272 Mb), 5 pair of chromosomes (2n=10), short generation time (~12 weeks), easy genetic transformation, self-fertility and simple growth requirements. The model plant properties and its close relationship to important crops, makes Brachypodium an attractive model plant for improvement of all temperate crop species, particularly the cereals. In recent years, in spite of an increasing interest in genomic and transcriptomic studies on Brachypodium, these are shown to be insufficient in understanding metabolic networks composed of biochemical reactions, mainly due to lack of correlation between mRNA and protein levels and the lack of enzymatic activity of proteins upon translation. Despite its importance of being the closest omic level to physiology, studies on metabolomics for Brachypodium are at their infancy. In order to elucidate the effect of drought stress on metabolism, the objective of this study is to set up quantitative metabolomics platform for Brachypodium distachyon, in particular, to optimize metabolite extraction protocols. For validation of extraction protocols, we used reporter metabolites (specifically ATP, glucose and overall starch) and focused on quantification of these, rather than overall, high-coverage metabolome data. As such, we evaluated the effect of drought stress and genotypic differences based on these reporter metabolites in model plant Brachypodium using two different genotypes (each represented by 2 populations). Careful analysis of reporter metabolites revealed that there is no “one-protocol-fits-all”, meaning that there is a significant difference in metabolite recoveries upon different extraction protocols. Moreover, the extraction efficiency is also affected by different genotypes, which resultingly calls for optimization of protocols, tailor-made for a genotype. Quantitative analysis of reporter metabolites points to significant changes upon drought stress, yet the resulting change differs from one metabolism to another, i.e. energy metabolism responds differently than glucose and carbohydrate metabolism. ATP levels exhibited (ranging from 2.92 10-5 – 1.77 10-3 nmol/g FW (fresh weight)) a different trend between the extraction protocols. Glucose levels showed 1-4 fold increase upon drought stress. Starch levels in Brachypodium leaves, ranging between 0.02-2.7% FW, exhibited no significant or consistent change, as a response to drought stress. On the basis of these results, it can be concluded that the impact of drought stress on Brachypodium metabolism was significant and this study could aid for further metabolomics studies on drought stress response for the improvement of agriculturally important crops.