Proteomics Study On Response Mechanism Of Heat Stress In Dahlia

Dahlia pinnata Cav.is originated in Mexican plateau area,which likes sunny,cool and well-ventilated environment.However,the high temperature and humid climate conditions in southern China seriously affect the normal growth and garden application of Dahlia.Studies have shown that exogenous salicylic acid can effectively increase the heat resistance of dahlia,but its stress response mechanism is not yet clear.Therefore,in this study,Dahlia 'dan ban huang' cultivars were used as materials,and proteomic methods were used to isolate and identify high temperature stress of Dahlia and exogenous salicylic acid-regulated response-related proteins.Molecular biology techniques were used to clone the heat shock protein gene,and analyze its expression characteristics to elucidate the molecular mechanism of the response to high temperature stress and the regulation of exogenous salicylic acid in Dahlia.The main experiment results are as follows:1.The petals and leaves of flowering plants in Dahlia are used as materials,368 and 456 proteins were successfully isolated and identified from petals and leaves by using SDS-PAGE and LC-MS/MS,respectively.By GO analysis,there were 356 and 451 proteins in the functional annotation of petals and leaves,which mainly including binding activity,catalytic activity,structural molecular activity,transporter activity,electron carrier activity,and antioxidant activity;there are 360 and 448 proteins in the petal and leaf biochemical annotations,which mainly involved in cellular processes,metabolic processes,single biological processes,biological regulation,stress response,and localization;there were 342 and 437 proteins with corresponding localization in petal and leaf components,which were mainly located in the cytoplasm,organelles,cell membranes,macromolecular complexes,and cell nucleus.2.The flowering of Dahlia be used as materials at a temperature of 35°C and with 25 °C as a control.The total protein of dahlia flower petals was extracted by phenol extraction method.50 significant expression spots?more than 2 times?were detected by use IPG/SDS-PAGE after silver staining and colorimetric analysis,of which 37 proteins were up-regulated,and 19 known functional proteins were successful identified,including 13 resistance associated proteins,2 cytoskeleton-forming proteins and 4 metabolism related proteins.3.Apply SA treatment?H₂O as control?to petals of Dahlia under high temperature stress at 35°C after 1 week,which the differential proteins were isolated and identified by using SDS-PAGE and Shotgun-LC/MS,and131,73,123,and 152 differential proteins were obtained in four gel bands.Applying bioinformatics analysis found that mainly includes the binding activity,catalytic activity,structural molecular activity,transporter activity,electron carrier,antioxidant activity;and it is mainly involved in biological processes such as cellular process,metabolic process,single-organism process,biological regulation,response to stimulus;and mainly distributed in organelles,cells,cell membranes,macromolecular complexes,septa,cell junctions.KEGG pathways mainly include photosynthesis,photosynthetic carbon fixation,glycolysis,glyoxylate and dicarboxylic acid metabolism,and nitrogen metabolism.4.The c DNA sequences of HSPs group genes were cloned from Dahlia petals by molecular biology and bioinformatics technologies,which were respectively named as DpHSP90?750bp,249 amino acid residues and one stop codon?,DpHSP70?705bp,234 amino acid residues and one stop codon?,DpCPN60?1569bp,522 amino acid residues and one stop codon?,DpHSP17?346bp,171 amino acid residues and one stop codon?.Homology analysis showed that the gene sequences of the members of the heat shock protein family in compositaes are fairly conservative.5.The HSPs group gene?DpHSP90,DpHSP70,DpCPN60,DpHSP17?of the Dahlia petals has been analyzed by q RT-PCR.The results show that four genes were expressed in petals,and showed the same tendency under high temperature stress,which was the highest expression in exogenous SA group with heat stress,followed by the high temperature stress group,and lowest in the normal temperature control group.These results indicate that the four heat shock proteins belong to the Dahlia response strain related to high temperature stress.Exogenous SA participates in the regulation of heat shock response,which can improve the plant's stress resistance and relieve the damage of high temperature stress to Dahlia.