【作者】 王哲；

【导师】 史国安；

【作者基本信息】 河南科技大学 ， 植物学， 2014， 硕士

【摘要】 牡丹在我国有“花王”的美誉，是我国特别珍贵的花卉品种。但是由于其花期短暂，现在已经难以满足人们的观赏需求。有研究表明MAPK级联途径与植物的生长发育有重要的联系，特别是和植物的开花衰老密切相关。MAPK级联途径是真核细胞中高度保守的信号通路，近年来，很多植物中的MAPKs的作用机理都有了长足进展，但关于牡丹开花衰老过程中MAPK信号转导方面的研究较少。本文以牡丹品种‘玉面桃花’为研究对象，在使用MAPK级联途径抑制剂PD98059和dH2O预处理12h后，分别在蒸馏水、0.1mmol/L ABA、10mmol/LH2O2、1mmol/L CEPA溶液中进行瓶插，通过测定瓶插过程中的形态和生理生化变化，以及ACS和ACO基因差异表达情况，对ABA、H2O2、CEPA以及MAPK信号通路在牡丹切花开放衰老进程中的生理效应进行了初步探索，以期为延长牡丹切花瓶插寿命和贮藏保鲜技术提供理论依据。现将结果报告如下：1. ABA、H2O2和CEPA处理加速了水分平衡值达到0值的时间，提高了开花速率，但缩短了牡丹切花的瓶插寿命，缩短了最佳观赏期，降低了牡丹切花的观赏品质，表明ABA、H2O2和CEPA具有促进牡丹切花衰老的作用。相比而言，CEPA促进衰老的作用较ABA和H2O2强。2.施用外源物质ABA、H2O2和CEPA处理后，增强了切花的呼吸速率，并且伴有明显的乙烯跃变，表明‘玉面桃花’为乙烯敏感型花卉。乙烯释放量的变化趋势与ACS活性变化趋势基本一致，乙烯合成与ACC含量以及ACO、ACS活性密切相关，进一步验证了ACC是乙烯生物合成过程中的直接前提物质，ACO、ACS是乙烯生物合成过程中的关键限速酶。MAPK级联途径抑制剂处理过后切花的呼吸速率及乙烯代谢都有所减弱，MAPK级联途径参与了切花的衰老调控，并参与到ABA、H2O2和CEPA的调控机制中。3. ABA、H2O2和CEPA处理一定程度上加速了MDA的积累，导致膜脂过氧化程度加剧，可溶性蛋白含量降低，抗氧化酶活性的增加都早于空白对照处理，并且抗氧化酶活性的变化与H2O2变化趋势基本一致。使用MAPK级联途径抑制剂PD98059处理后，MDA含量、可溶性蛋白与抗氧化酶活性与对照相比，均有所降低，表明MAPK级联途径参与了牡丹切花开花衰老过程中的信号传导。4.半定量RT-PCR分析显示牡丹Ubiquitin基因在不同处理不同发育时期的表达量相对恒定，可作为半定量分析的内参基因。对照的ACO和ACS基因基本表现为在瓶插初期表达量略大，瓶插中期表达量有所下降，推测可能是ACO和ACS活性比较高，反馈调节抑制了基因表达。在瓶插后期，即盛花期至始衰期，基因表达量又有所增加。使用ABA、H2O2、CEPA及MAPK级联途径抑制剂后，ACO和ACS基因表达量较对照有所差异，表明MAPK级联途径参与到ABA、H2O2、CEPA调控衰老的进程中。更多还原

【Abstract】 As the king of flower, peony is an especially precious flower variety in ourcountry. Now it is hard to meet peoples’ needs because of its short flowering. Studieshave shown that MAPK play an important role in plants’ growth. especially for plantsblossom and aging. MAPK cascade was highly conserved modules in all eukaryote. Inrecent years, numerous MAPKs had been isolated from different plants, and studiedthe function of MAPKs. But the relevant research on peony was infrequent.Peony (Paeonia suffruticosa ‘Yumiantaohua’) cut flowers were taken as objectsof study. They were divided into two groups, one pretreated by0.1mM PD98059(MAPK specific inhibitor) for12h, another one pretreated by ditilled water. Eachgroups were divided into four equal groups and randomly put in distilled water,0.1mM ABA,10mM H2O2and1mM CEPA respectively for analysing the morphologicalof flower braches and physiological and biochemical indexes during vasing. Throughstudying analyse the physiological effect of ABA, H2O2, ethylene and MAPKsignaling passway on the aging process of peony cut flower provide theoretical basisfor fresh-keeping technology and prolonging its vase days. The main results were asfollows:1. ABA、H2O2and CEPA accelerated the time of water balance, shortened theperiod of peony flowering, promoted flower opening and increased the maximumflower diameter. The treatment also shortened the best viewing period, and reduced thepeony ornamental quality of cut flowers. The result showed that ABA、H2O2andCEPA could promote flower opening and senescence. CEPA was the strongest factor topromote flower opening and senescence, the influence of ABA and CEPA wererelatively weak.2. Exogenous ABA、H2O2and CEPA could enhance the respiratory rate offlowers, and accompanied ethylene jump phenomenon. It showed that ‘Yumiantaohua’ was ethylene sensitive flower. The change trend of ethylene release was similar to theACC content and the activity of ACO、ACS. It meant that ACC was the premisematerial in the process of ethylene compound, and the ACO、ACS were the importantenzyme of speed in the process of ethylene compound. MAPK cascade pathwaysinvolved the aging of cut flower, and also participated in the regulatory mechanism ofABA、H2O2and CEPA, in addition MAPK cascade pathways had the positive controlin this process.3. ABA、H2O2and CEPA treatments could accelerate the accumulation of MDA,increase membrane-lipid peroxidation, reduce soluble protein content. The activity ofantioxidant enzyme increased earlier than blank treatment, and its change trend in thesame way as H2O2. After using the PD98059, the MDA content、soluble protein andactivity of antioxidant enzymes were reduced compared with the blank control.Therefore, MAPK maybe play an important role in the pathway of the cut peonyflowers senescence.4. Smi-quantitative RT-PCR analysis showed that the gene expression leves ofUbiquitin gene in different developmental stages and different processing of peonywere almost constant. The ACO and ACS genes at the beginning of the vase to expressquantity slightly larger, vase and mid expression quantity decreased, That may becoated the ACO and ACS activity is higher, feedback regulation inhibit geneexpression.Beginning in the late vase, namely into decayed, gene expression quantityincreased again. AS the contrast, the amount of gene expression of ACS and ACO waslarge at the beginning of the vase. Then it was decreased in the middle of the vase.This was likely to be the activity of ACS and ACO was higher, made the feedbackcontrol. But late in the vase, the amount of gene expression was increase. After usedABA、H2O2、CEPA and PD98059, the gene expression of ACS and ACO wasdifferent from the contrast, this showed that the MAPK cascade pathway involved inthe pathway of ABA、H2O2and CEPA regulated flowers senescence.更多还原