Studies On Mechanisms Of Salt Tolerance In Sea Barley And Functional Characterization Of HKT1;5 In Barley

Soil salinization is a major abiotic stress restricting sustainable agricultural development worldwide.Understanding of physiological and molecular mechanisms of salt tolerance in crops is quite imperative for breeding of salt-tolerant crop varieties.Barley(Hordeum vulgare)is a cereal crop with high salt tolerance.Tibetan annual wild barley(Hordeum vulgare ssp.spontaneum)is rich in genetic diversity,containing elite accessions with stronger salt tolerance than cultivated barley.In addition,sea barley(Hordeum marinum)has been shown to be much more salt tolerance than barley.In this thesis,we compared the differences in salt stress response among cultivated barley,wild barley and sea barley in terms of physiological phenotype,metabolome and transcriptome changes,and had an initial understanding of the mechanisms for the salt tolerance of sea barley.Then we studied the biological functions of HvHKTl;5,a key gene of salt stress response from barley,and its homologous gene in sea barley,HmHKT1;5.The molecular mechanisms of the difference in salt tolerance between barley and sea barley were analyzed.The main results of this study are as follows:1.Understanding of the superior salt tolerance and its physiological mechanism in sea barleySea barley H559 showed much higher salt tolerance than XZ113 and Golden Promise,and its mechanisms for salt tolerance could be summarized as follows:(1)maintaining ion balance by regulating the expression of the genes related to Na+ and K+ transport;(2)taking less energy-consuming strategies,including a larger scale gene down-regulation and utilization of inorganic ions(i.e.Na+ and K+)as "cheap"osmotica for root tolerance;(3)enhancing glycolysis and the TCA cycle for energy supply and compatible solute accumulation for shoot tolerance.2.Determination of the gene structure and expression pattern of HvHKTl;5HvHKT1;5 was obtained by homologous cloning from a cultivated barley Golden Promise,and it had the highest similarity with TaHKT1;5-B2 in protein sequence.There are three specific amino acid residues in PA and PB of HvHKT1;5 compared with rice and wheat homologous HKT1;5s.HvHKT1;5 was mainly expressed in roots,being up-regulated by salt stress.HvHKT1;5 is only permeable to Na+,and its transport activity is inhibited by external K+.3.Understanding of the mechanisms of HvHKTl;5 for salt tolerance.HvHKTl;5 RNAi and over-expression transgenic lines were obtained using Gateway and barley transformation approaches.The salt tolerance of those transgenic lines was evaluated.The results showed that knock-down of HvHKT1;5 reduced Na+uptake and translocation,resulting in the enhanced salt tolerance,and overexpression of HvHKT1;5 caused more salt sensitivity.Unlike HKT1;5s in other gramineous crops,HvHKT1;5 mediates Na+ uptake in root epidermis and promotes Na+ translocation from roots to shoots through xylem,negatively regulating salt tolerance in barley plants.4.Revealing of the functions of barley and sea barley HKT1;5 transporters and their differences in expressionThe similarity of amino acid sequence for HKT1;5s of barley and sea barley was as high as 93%,and the gene expression patterns were quite similar in term of the expressed tissue and salt-induced up-regulation,but the size of effect was different.HmHKT1;5 was permeable to Na+,similar to HvHKT1;5,but the ion affinity was much lower than HvHKT1;5 in X.laevis oocytes.Both HmHKTl;5 and HvHKT1;5 were involved in Na+ uptake and Na+ loading from roots to shoots,but the former showed much weaker transport ability.The current results indicate that the relatively lower expression level of HmHKT1;5,as well as its weaker Na+ affinity,Na+ uptake and translocation ability could be a major reason for the higher salt tolerance of sea barley in comparison with barley.