Analysis Of Proteome Of The Alfalfa Seedling Under Salt Stress And Cloning Of The Salt-stress Related Genes

The present study is that the response of alfalfa seedlings to salt stress during germinationwas investigated at both physiological and proteomic levels. The difference of satl tolerencebetween Zhongmu NO.3and Daxiyang was determinated in terms of physiological level.Furthermore, based on the two dimensional electrophoresis, the protein expression patterns ofalfalfa seedlings from both of the cultivars were compared when they grew under200mmol/L for7days. A great number of differently expressed proteins under salt stress were successfullyidentified in our research. A new gene, named annexin, was chosen to clone based on a dramaticdown-regulation in salt-sensitive cultivar while no significant change in salt-tolerant in theprotein abundance. And its functions were further explored in present research. All in all,ourresearch provides basic data for analyzing the molecular mechanisms of salt stress response. Thedetailed research results are as follows:1. The germination rate of four alfalfa cultivars was comparatively studied under0,100,150,200mmol/L NaCl treatment. Alfalfa Zhongmu NO.3and Daxiyang varieties were selected asstress-tolerant and-sensitive cultivars, respectively, and were used for furthercharacterization.2. The difference of alfalfa seedlings in terms of salt tolerance between the cultivars wereanalysized via some physiological items such as biomass, proline content, chlorophyllcontent.3. In according with comparison and statistical analysis, it was found that91protein spots hadsignificant changes in abundance(p<0.05). Among them,20sports were up-regulated while71sports were down-regulated under salt stress.36proteins were identified from these91spots by MALDI-TOF/TOF-MS.4. According to analysis of function of identified proteins, five identified proteins includedrubisco activase, fructokinase, dihydroflavonol4-reductase, alcohol dehydrogenase,annexin.They are involved in photosynthesis, energy metabolisms,secondary metabolisms,detoxifying.5. An identified protein annexin was determianted as a novel salt-tolerance-associated protein.According to the information of the protein, the OFR(open reading frame) of new genenamed MsAN was cloned, which contained945bp and encode the protein made of314aminoacids. The protein included4unique annexin domains.6. In transcriptional level, the expression of MsAN was induced by NaCl, ABA, PEGandH2O2,indicating that MsAN may be involved in some abiotic stress responses in alfalfa.