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两种彩叶植物叶色表达相关机理研究

Studies on the Leaf Color Expression Related Mechanism for Two Colorful Plants

作者: 专业:园林植物与观赏园艺 导师:苑兆和 年度:2010 学位:硕士  院校: 山东农业大学

Keywords

Cercis canadensis‘Forest Pansy’, Populus×euramericana‘Zhonghong’, anthocyanin, solubility sugar, chlorophyll, PAL, CHI, DFR, UFGT

        本研究以‘紫叶加拿大紫荆’(Cercis canadensis‘Forest Pansy’)和‘中华红叶杨’(Populus×euramericana‘Zhonghong’)为试材,通过对其叶片色素合成动态变化、相关物质动态变化、酶活性的研究以及‘紫叶加拿大紫荆’CHI基因克隆初步研究,对这两种彩叶树种叶片呈色的生理机制进行了初步探讨,并得出以下主要结论:(1)‘紫叶加拿大紫荆’和‘中华红叶杨’两种彩叶植物叶色表达期叶片中色素的总体变化规律相似。‘紫叶加拿大紫荆’叶片呈色期较长,从4月中旬~6月初。5月初气温较低,花青苷含量相对较高,5月底~6月初气温迅速升高,花青苷的含量下降,总叶绿素和类胡萝卜素含量迅速升高;而’中华红叶杨’的叶片呈色期较短,从4月中旬~5月上旬。‘紫叶加拿大紫荆’整个叶色表达期花青苷呈总体下降的“M”型趋势,‘中华红叶杨’花青苷含量呈升-降-升-降的变化趋势。(2)‘紫叶加拿大紫荆’和‘中华红叶杨’叶片颜色的变化主要是花青苷降解与叶绿素合成的结果,叶片的呈色取决于花青苷和总叶绿素的比例。当花青苷与总叶绿素的比例小于1:0.18时,‘紫叶加拿大紫荆’叶片开始由鲜亮的紫红色向暗红色转变,当比例小于1:1.92左右时叶片由暗红色转变为绿色;在‘中华红叶杨’中,当花青苷与总叶绿素的比例小于1:0.66时叶片开始由亮紫红色向暗红色转变,当比例小于1:3.23时叶片由暗红色转向绿色。(3)对‘紫叶加拿大紫荆’和‘中华红叶杨’叶色表达期叶片进行花青苷、叶绿素、类胡萝卜素和可溶性糖的测定,并进行相关性分析。结果表明,‘紫叶加拿大紫荆’叶片中类胡萝卜素、总叶绿素与花青苷均呈极显著的负相关,随着花青苷含量的降低,叶绿素和类胡萝卜素含量不断升高;可溶性糖与紫叶加拿大紫荆叶片中花青苷含量的相关性不显著。‘中华红叶杨’中,总叶绿素、类胡萝卜素与花青苷呈显著的负相关关系,可溶性糖含量与花青苷存在极显著的正相关关系,随着可溶性糖含量的降低,花青苷含量降低。(4)测定了两种彩叶树种叶色表达期叶片中花青苷含量,以及花青苷合成相关酶苯丙氨酸解氨酶(PAL)、查尔酮异构酶(CHI)、二氢黄酮醇还原酶(DFR)、类黄酮糖基转移酶(UFGT)活性的变化。结果表明,4月中旬~6月初,‘紫叶加拿大紫荆’叶片中花青苷含量下降了93%,‘中华红叶杨’叶片中花青苷含量下降81%。‘紫叶加拿大紫荆’叶色表达期,PAL酶活性与花青苷的含量呈三次曲线关系,CHI酶活性与花青苷的含量呈极显著的倒数曲线关系,随着CHI酶活性的升高,花青苷含量降低。DFR酶活性与花青苷的含量呈极显著的线性关系,随着DFR酶活性的升高,花青苷含量增加,随着DFR酶活性的降低,花青苷含量降低,UFGT酶活性与花青苷含量呈指数曲线关系;‘中华红叶杨’叶片转色期花青苷合成相关酶的活性变化研究结果表明,PAL酶活性与花青苷的含量呈二次曲线关系,当PAL酶活性小于625.84U·g-1FW时,花青苷含量随着PAL酶活性的降低而降低。CHI酶活性与花青苷含量存在“S”曲线关系,DFR酶活性与花青苷含量存在三次曲线关系,UFGT与花青苷的合成存在极显著的指数曲线关系,随着UFGT酶活性的降低,花青苷含量降低,UFGT酶活性较高时,花青苷含量也较高,因此提高其活性能提高‘中华红叶杨’叶片中花青苷的含量。
    The synthetic dynamic changes of pigments, related substances and enzyme activity of leaf color expression were studied in Cercis canadensis‘Forest Pansy’and Populus×euramericana‘Zhonghong’. The preliminary study of CHI gene cloning was carried on the Cercis canadensis‘Forest Pansy’. The physiological mechanisms of leaf colouration in two colorful plants were discussed. The main results were submitted as follows:(1) The change regular of pigment in two colorful plants were similar during leaf color expression. The period of leaf color expression in Cercis canadensis‘Forest Pansy’was much longer, which lasted from the middle April to early June. The content of anthocyanin was relatively higer in early May, because temperature was much lower. Total chlorophyll and carotenoid content increased rapidly while anthocyanin content decreased from late May to early June. The period of leaf color expression in Populus×euramericana‘Zhonghong’was short ,which lasted from middle April to early May. The anthocyanin content decreased and presented“M”tendency with the days increased in Cercis canadensis‘Forest Pansy’. The change tendency of anthocyanin content increased first and descended later in Populus×euramericana cv. Zhonghuahongye.(2) The color change of leaves was the result of anthocyanin degradation and chlorophyll synthesis in two colorful plants. The leaf color expression was determined by the ratio of anthocyanin and total chlorophyll. For Cercis canadensis‘Forest Pansy’, the leaves color turned dark red when the ratio of anthocyanin and total chlorophyll was less than 1:0.18, the leaves color turn green when the ratio of anthocyanin and total chlorophyll was less than 1:1.92. For Populus×euramericana‘Zhonghong’, the leaves color turn dark red when the ratio of anthocyanin and total chlorophyll was less than 1:0.66, the leaves color turn green when the ratio of anthocyanin and total chlorophyll was less than 1:3.23.(3) The content of pigments and other chemical components includinganthocyanin,chlorophyll,carotenoids and solubility sugar were investigated and analysedtheir correlation in two colorful plants,. The result indicated that the cholorphyll andcarotenoid had remarkablely negative correlation with the anthocyanin in Cerciscanadensis‘Forest Pansy’. The soluble sugar was not correlated with the anthocyanin inCercis canadensis‘Forest Pansy’. The correlation of cholorphyll ,carotenoid andanthocyanin were significant in Populus×euramericana‘Zhonghong’. The correlationof soluble sugar and anthocyanin were remarkablely positive in Populus×euramericana cv. Zhonghuahongye. The content of soluble sugar decreased with theanthocyanin content reducing.(4) Content of anthocyanin and the activities of enzymes in anthocyaninbiosyntheses including plenylalanine ammonia lyase (PAL), chalcone ismoerase (CHI),dihydroflavonol reductase (DFR) and UDP glucose-flavonoid-3-o-glycosyltranfe-rase(UFGT) were investigated in two colorful plants during leaf color expression. The resultsshowed that the content of anthocyanin in Cercis canadensis‘Forest Pansy’decreased by93% from the middle April to early June. The content of anthocyanin in Populus×euramericana‘Zhonghong’decreased by 81%. During the period of leaves colorexpression of Cercis canadensis‘Forest Pansy’, the relationship of PAL activity andanthocyanin content presented a cubic curve . The activity of CHI was significantlycorrelated with the content of anthocyanin. when the content of anthocyanin decreased,the activity of CHI increasing. The activity of DFR had a remarkable linear relationshipwith the content of anthocyanin. When the content of anthocyanin decreased, the activityof DFR reduced. The activity of UFGT had a exponential curve relationship with thecontent of anthocyanin. For Populus×euramericana‘Zhonghong’, the correlation between PAL activity and anthocyanin presented a quadratic curve . When the activity of PAL was below 625.84U·g-1FW, the content of anthocyanin decreased with the activity of PAL reduced. The relationship of CHI activity and anthocyanin content had a‘S’curve. The activity of DFR and anthocyanin content had a cubic curve. There was a extremely significant exponential curve relationship between the activity of UFGT and anthocyanin. The content of anthocyanin became high as the activity of UFGT was high, which became low as the activity was low. To increase the activity of UFGT could improve the content of anthocyanin in Populus×euramericana‘Zhonghong’.
        

两种彩叶植物叶色表达相关机理研究

摘要8-10
ABSTRACT10-12
1 引言13-30
    1.1 彩叶植物的研究进展13-18
        1.1.1 彩叶植物的定义与分类13
        1.1.2 彩叶树叶片中色素的研究13-14
        1.1.3 彩叶植物色彩发育过程中的形态解剖研究14-15
        1.1.4 彩叶植物叶片呈色机理15-16
        1.1.5 环境因子对叶片呈色的影响16-17
        1.1.6 彩叶植物的应用17-18
    1.2 花青苷及生物合成18-26
        1.2.1 花青苷概述18-19
        1.2.2 花青苷生物合成途径19-20
        1.2.3 花青苷合成与糖的代谢20-21
        1.2.4 花青苷合成过程中酶的调节21-23
        1.2.5 花青苷合成过程中关键基因的克隆23-26
    1.3 花青苷对植物适应环境的意义26-28
        1.3.1 提高光保护能力26
        1.3.2 提高抗冻能力26-27
        1.3.3 提高抗旱能力27
        1.3.4 提高抗氧化能力27
        1.3.5 提高抗菌能力27-28
    1.4 彩叶植物研究展望28
    1.5 紫叶加拿大紫荆和中华红叶杨的概况28-29
    1.6 本研究的目的与意义29-30
2 试验材料与方法30-36
    2.1 两种彩叶植物叶色表达期相关物质的动态变化30
        2.1.1 试验材料30
        2.1.2 色素含量的测定30
        2.1.3 可溶性糖含量测定30
    2.2 两种彩叶植物叶色表达期花青苷合成相关酶的活性变化30-32
        2.2.1 试验材料30-31
        2.2.2 PAL 和 CHI 酶活性的测定31
        2.2.3 DFR 和 UFGT 酶活性的测定31-32
    2.3 紫叶加拿大紫荆叶片中花青苷合成关键酶基因 CHI 克隆的初步研究32-35
        2.3.1 试验材料32
        2.3.2 叶片RNA 的提取方法32-34
        2.3.3 反转录34
        2.3.4 PCR 反应34-35
        2.3.5 目的片段回收35
    2.4 数据统计分析35-36
3 结果与分析36-51
    3.1 两种彩叶植物叶色表达期相关物质的动态变化36-44
        3.1.1 色素含量动态变化36-42
        3.1.2 可溶性糖含量变化动态42-43
        3.1.3 叶绿素、类胡萝卜素、可溶性糖含量与花青苷的相关分析43-44
    3.2 两种彩叶植物花青苷合成相关酶的活性变化44-49
        3.2.1 紫叶加拿大紫荆叶色表达期相关酶活性动态变化44-45
        3.2.2 中华红叶杨叶色表达期相关酶活性动态变化45-48
        3.2.3 两种彩叶植物花青苷含量及其相关酶活性的相关性分析48-49
    3.3 紫叶加拿大紫荆叶片CHI 基因克隆的初步研究49-51
        3.3.1 叶片RNA 的提取49-50
        3.3.2 RT-PCR50-51
        3.3.3 目的片段回收51
4 讨论51-56
    4.1 两种彩叶植物叶色表达期色素含量的动态变化51-52
    4.2 两种彩叶植物叶色表达期相关物质的相关性52-53
    4.3 两种彩叶植物叶色表达期花青苷合成相关酶的活性变化53-55
    4.4 紫叶加拿大紫荆花青苷合成关键基因 CHI 克隆的初步研究55-56
5 结论56-58
参考文献58-71
致谢71-72
攻读硕士学位期间发表的论文72
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