洋桔梗切花衰老过程中的生理生化分析
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摘要
洋桔梗(Eustoma grandiflorm(Raf.)Shnn)，是龙胆科草原龙胆属观赏切花，其花色淡雅、花姿优美，市场开发潜力巨大，已成为又一切花新秀。洋桔梗由于采后贮运及瓶插过程中容易发生花朵不能正常开放、花瓣过早萎蔫、花枝折断、花茎弯曲、茎叶黄化、干枯等问题，影响了切花的观赏价值。本研究以洋桔梗品种“白金紫”、“国王粉”和“罗莎绿”为研究材料，用乙烯促进剂乙烯利和乙烯拮抗剂1-MCP对洋桔梗进行处理，研究洋桔梗花瓣衰老过程中水分含量、蛋白质含量、保护酶活性等生理变化情况，并应用SDS-PAGE电泳技术对花瓣中蛋白质变化情况进行研究，以期从分子水平上研究洋桔梗切花衰老过程，并能为洋桔梗的遗传育种提供理论依据。同时以白金紫为试材，从细胞学角度研究了洋桔梗的衰老过程。
本试验研究结果如下：
1、用浓度分别为25mg／L、50 mg／L、100 mg／L、200 mg／L、400 mg／L的乙烯利24h和浓度分别为500mg／L(8h、10h)、800mg／L(8h、10h)的1-MCP处理白金紫的花枝，结果表明：用200mg／L的乙烯利处理24h明显抑制白金紫花瓣正常生长发育，缩短花瓣的寿命，降低洋桔梗的观赏品质；800 mg／L的1-MCP处理8h可以较为理想地延缓白金紫花瓣的衰老，延长瓶插寿命，提高观赏价值。
用浓度为200mg／L的乙烯利处理白金紫、国王粉、罗莎绿花瓣24h后，含水量下降较快，抑制了保护酶的活性，可溶性糖和可溶性蛋白质的含量也下降较快，明显的促进了洋桔梗花瓣的衰老。用800mg／L的1-MCP处理8h后的白金紫、国王粉、罗莎绿花瓣，含水量下降较慢，保护酶活性强，可溶性总糖和可溶性蛋白质含量增加，对洋桔梗起到了很好的保鲜延衰作用。
2、通过SDS-PAGE凝胶电泳以及计算机辅助的图像分析技术，以白金紫、国王粉、罗莎绿三个品种作为试验材料，用清水、800mg／L的1-MCP处理8h和200mg／L的乙烯利处理24h，得到三种处理下的蛋白质图谱。分析结果表明：与对照相比，1-MCP处理的蛋白谱带多而浓，乙烯利处理的蛋白谱带急剧减少或是变淡。蛋白质谱带的变化有四种情况：有的谱带基本保持稳定不变；有的谱带减少消失；有的新合成的谱带出现后很快消失；有的新谱带出现后较为稳定。
3、三个洋桔梗切花品种对乙烯利、清水和1-MCP作用的敏感性不同，但同一药剂对三个品种的作用表现出相似的变化规律。
4、通过透射电镜对白金紫花瓣三个时期即绿蕾期、盛花期和衰败期的超微结构进行观察，结果表明：白金紫花瓣的衰老具有植物细胞程序性死亡的特点，如细胞核和细胞质凝缩、染色质边缘化、细胞和细胞器的空腔化、凋亡小体形成等；更表现为植物细胞在逆境下的受诱导程序性死亡特点，如发生质壁分离、出现液泡早早就消亡等水分胁迫下的特点；还观察到在鲜花怒放的盛期花瓣内有大量的淀粉粒和脂肪滴，绿蕾期和衰败期则没有，所以可以认为洋桔梗花瓣是在盛花期为以后的代谢贮存了主要的能量物质，这可能也是洋桔梗切花具有较长的瓶插寿命的一个重要原因。
Abstract:
Eustoma grandiflorm, belonging to Eustoma ressellianum, gentianaceae, hasbecome a promising flower in the cut-flower market due to its elegant color andbeautiful appearance. However, such problems as abnormal flowering, prematurepetal wilting, branch break, scape bending, haulm yellowing or withering often occurin the post-harvest transport and during the vase-holding process, seriously affectingthe ornamental value. We focused on the physiological changes, including watercontent, soluble sugar content, soluble protein content, activity of POD and activity ofSOD during the vase-holding process after the Eustoma grandiflorm var. of PlatinaViolet, King Pink and Rose Green were treated by using ETP and 1-MCP. Also, thechanging of petal protein was further studied by means of SDS-PAGE electrophoretictechnique with a view of deeply studying the senescence of cut-Eustoma at molecularlevel and provide some theoretical reference for Genetic Breeding. Meantime, takingthe King Pink as experimental material, the senescence process of cut-Eustoma is alsostudied in the context of cytology.
The result as follows:
1．ETP concentration(24h), respectively 25mg/L、50mg/L、100mg/L、200mg/L、400 mg/ L, and 1-MCP concentration, respectively 500mg/L (8h、10h)、800mg/L (8h、10h), were used for treating flower branch of Platina Violet petal. The result showedthat 200mg/L ETP for 24h is capable of distinctly inhibiting the normal developmentof Platina Violet petal, shortening the petal life, and reducing its ornamental quality.800mg/L1-MCP for 8h more ideally enables Platina Violet petal to postpone senility,extend the vase life, and improve the ornamental value.
2．By SDS-PAGE gel. electrophoresis technique and computer-aided image analysis,three varieties of Platina Violet, King Pink and Rose Green were taken asexperimental material, and then treated by using distilled water, 800mg/L 1-MCP for8h and 200mg/L ETP for 24h. By doing so, three types of protein maps were obtained.The result showed that in contrast with the control, 1-MCP lanes were more andthicker than water lanes; ETP lanes were fewer and thinner than water lanes. As aresult, there were four change trends: some lanes was relatively steady, some lanesdisappeared, some lanes flashed and some lanes stayed for a long time after composed
3．Though Platina Violet, King Pink and Rose Green had different reactivity after treatments of ETP、water and 1-MCP, they had similar change trend in sametreatment.
4．The ultra-structure during bud, blooming and wilting period of Platina Violet petalwere observed by means of TEM (transmission electron microscopy). The resultshowed that the senescence of cut-Eustoma petal was a typically programmed celldeath, for example, karyon and nucleolus and cytoplasm concentrating, chromatinedging, cell and organelle cavuming, languish-ball appearing etc. Further, it wascharacterized as the induced programmed cell death under adverse circumstance,including plasmolysis, vacuole dying out too early. In addition, lots of fecula and fatballs were found during blooming period, but not during bud and wilting periods.Therefore, it can be concluded that Eustoma petal accumulated the main energymaterial for future metabolism during the booming period. This may be one of the keyreasons that cut-Eustoma has a longer vase life.
本试验研究结果如下：
1、用浓度分别为25mg／L、50 mg／L、100 mg／L、200 mg／L、400 mg／L的乙烯利24h和浓度分别为500mg／L(8h、10h)、800mg／L(8h、10h)的1-MCP处理白金紫的花枝，结果表明：用200mg／L的乙烯利处理24h明显抑制白金紫花瓣正常生长发育，缩短花瓣的寿命，降低洋桔梗的观赏品质；800 mg／L的1-MCP处理8h可以较为理想地延缓白金紫花瓣的衰老，延长瓶插寿命，提高观赏价值。
用浓度为200mg／L的乙烯利处理白金紫、国王粉、罗莎绿花瓣24h后，含水量下降较快，抑制了保护酶的活性，可溶性糖和可溶性蛋白质的含量也下降较快，明显的促进了洋桔梗花瓣的衰老。用800mg／L的1-MCP处理8h后的白金紫、国王粉、罗莎绿花瓣，含水量下降较慢，保护酶活性强，可溶性总糖和可溶性蛋白质含量增加，对洋桔梗起到了很好的保鲜延衰作用。
2、通过SDS-PAGE凝胶电泳以及计算机辅助的图像分析技术，以白金紫、国王粉、罗莎绿三个品种作为试验材料，用清水、800mg／L的1-MCP处理8h和200mg／L的乙烯利处理24h，得到三种处理下的蛋白质图谱。分析结果表明：与对照相比，1-MCP处理的蛋白谱带多而浓，乙烯利处理的蛋白谱带急剧减少或是变淡。蛋白质谱带的变化有四种情况：有的谱带基本保持稳定不变；有的谱带减少消失；有的新合成的谱带出现后很快消失；有的新谱带出现后较为稳定。
3、三个洋桔梗切花品种对乙烯利、清水和1-MCP作用的敏感性不同，但同一药剂对三个品种的作用表现出相似的变化规律。
4、通过透射电镜对白金紫花瓣三个时期即绿蕾期、盛花期和衰败期的超微结构进行观察，结果表明：白金紫花瓣的衰老具有植物细胞程序性死亡的特点，如细胞核和细胞质凝缩、染色质边缘化、细胞和细胞器的空腔化、凋亡小体形成等；更表现为植物细胞在逆境下的受诱导程序性死亡特点，如发生质壁分离、出现液泡早早就消亡等水分胁迫下的特点；还观察到在鲜花怒放的盛期花瓣内有大量的淀粉粒和脂肪滴，绿蕾期和衰败期则没有，所以可以认为洋桔梗花瓣是在盛花期为以后的代谢贮存了主要的能量物质，这可能也是洋桔梗切花具有较长的瓶插寿命的一个重要原因。
Abstract:
Eustoma grandiflorm, belonging to Eustoma ressellianum, gentianaceae, hasbecome a promising flower in the cut-flower market due to its elegant color andbeautiful appearance. However, such problems as abnormal flowering, prematurepetal wilting, branch break, scape bending, haulm yellowing or withering often occurin the post-harvest transport and during the vase-holding process, seriously affectingthe ornamental value. We focused on the physiological changes, including watercontent, soluble sugar content, soluble protein content, activity of POD and activity ofSOD during the vase-holding process after the Eustoma grandiflorm var. of PlatinaViolet, King Pink and Rose Green were treated by using ETP and 1-MCP. Also, thechanging of petal protein was further studied by means of SDS-PAGE electrophoretictechnique with a view of deeply studying the senescence of cut-Eustoma at molecularlevel and provide some theoretical reference for Genetic Breeding. Meantime, takingthe King Pink as experimental material, the senescence process of cut-Eustoma is alsostudied in the context of cytology.
The result as follows:
1．ETP concentration(24h), respectively 25mg/L、50mg/L、100mg/L、200mg/L、400 mg/ L, and 1-MCP concentration, respectively 500mg/L (8h、10h)、800mg/L (8h、10h), were used for treating flower branch of Platina Violet petal. The result showedthat 200mg/L ETP for 24h is capable of distinctly inhibiting the normal developmentof Platina Violet petal, shortening the petal life, and reducing its ornamental quality.800mg/L1-MCP for 8h more ideally enables Platina Violet petal to postpone senility,extend the vase life, and improve the ornamental value.
2．By SDS-PAGE gel. electrophoresis technique and computer-aided image analysis,three varieties of Platina Violet, King Pink and Rose Green were taken asexperimental material, and then treated by using distilled water, 800mg/L 1-MCP for8h and 200mg/L ETP for 24h. By doing so, three types of protein maps were obtained.The result showed that in contrast with the control, 1-MCP lanes were more andthicker than water lanes; ETP lanes were fewer and thinner than water lanes. As aresult, there were four change trends: some lanes was relatively steady, some lanesdisappeared, some lanes flashed and some lanes stayed for a long time after composed
3．Though Platina Violet, King Pink and Rose Green had different reactivity after treatments of ETP、water and 1-MCP, they had similar change trend in sametreatment.
4．The ultra-structure during bud, blooming and wilting period of Platina Violet petalwere observed by means of TEM (transmission electron microscopy). The resultshowed that the senescence of cut-Eustoma petal was a typically programmed celldeath, for example, karyon and nucleolus and cytoplasm concentrating, chromatinedging, cell and organelle cavuming, languish-ball appearing etc. Further, it wascharacterized as the induced programmed cell death under adverse circumstance,including plasmolysis, vacuole dying out too early. In addition, lots of fecula and fatballs were found during blooming period, but not during bud and wilting periods.Therefore, it can be concluded that Eustoma petal accumulated the main energymaterial for future metabolism during the booming period. This may be one of the keyreasons that cut-Eustoma has a longer vase life.
引文
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