Inducing salt tolerance in rice (Oryza sativa L.) varieties by gamma radiation

In context of this Ph. D. thesis study, it is aimed to obtain salinity tolerance on Osmancık-97, which is widely planted rice variety in our country, and Mevlütbey, which is a rice variety improved in Black Sea region, by using gamma radiation. Rice seeds were irradiated by 0, 250, 300 ve 400 Gy dose of Cs-137 source. Irradiated seeds were planted in the experimental fields of Black Sea Agricultural Research Institute and physiological effects of radiation on M1 generation were observed, as well as seed were harvested. Selections in M2 generation for determination of salt tolerant mutants were conducted in Plant Biotechnology Laboratory of T.C. Istanbul Kultur University, Department of Molecular Biology and Genetics. Following in vivo selections, 43 of selected salt tolerant mutants were belonging to Osmancık-97 variety. Selected M2 generation mutant plants were planted to Trakya Agricultural Research Institute to obtain seeds. In vitro callus cultures were established in order to determine persistence of salt tolerance in M3 generation. Embrios of the seeds were selected as explant source. B3 medium (MS salts, 3 mg/L 2,4-D, 560 mg/L proline) is determined to be the most suitable medium for callus induction. In respect to the data obtained from in vitro studies, 250-2, 300-5, 400-2 and 400-5 mutants which presented better callus growth comparing to the untreated Osmancık-97 control calluses were selected for further molecular characterization studies. Biochemical responses of selected salt tolerant mutants under salt treated and untreated conditions were evaluated by lipid peroxidation, proline content, photosynthetic pigment content, total protein content, enzyme activities of peroxidase, superoxide dismutase and catalase results. Genetic distance between mutants and Osmancık-97 as their origin were determined by PCR reactions conducted with ISSR primers. Jaccard genetic similarity matrix was generated according to the band profiles and UPGMA dendrogram was drawn. According to the dendrogram, Osmancık-97 variety and salt treated/untreated mutants divided into two distinct clusters. Moreover, all the mutants were seperated from Osmancık-97 variety in different sub-clusters. Two dimensional proteomic analysis of salt tolerant mutants were conducted. Evaluation of protein spot profiles of samples presented that 30 proteins were differed in expression. 9 of these 30 proteins (ribulose biphosphate carboxylase/oxidase activase, triose phosphate isomerase, 2-Cys peroxiredoxin, oxygene-evolving enhancer protein 1, ferritine-1, ferredoxin-NADP reductase, glyceraldehyde 3-phosphate dehydrogenase, tubby-like F-box protein 11, peptydil-prolyl cis-trans isomerase) were characterized by mass spectroscopy and they were found to be salt tolerans related. In this study, proteomic response differences between rice mutants under salt stress were found to be related to the roles of ferredoxin-NADP reductase, oxygene-evolving enhancer protein 1, triose phosphate isomerase and 2-Cys peroxiredoxin proteins. As a result, it is the first time in Turkey that we presented salt tolerant rice mutants can be induced by gamma radiation. In this study, four different salt tolerant rice mutants were obtained. These four mutants will play important roles on improving novel rice varieties which have higher yield potential under different environmental, climate and geographical conditions. Also, this study is important for our country's economy due to the fact that our salt tolerant mutants were induced from our local rice varieties.