【作者】 徐幸福；

【导师】 王彩云；

【作者基本信息】 华中农业大学 ， 园林植物与观赏园艺， 2003， 硕士

【摘要】 桂花(Osmanthus fragrans Lour)由于其特有的芳香而极具观赏性，在园林栽培中广泛运用。随着大花品种的出现，桂花已经成为一种极具发展潜力的切花资源，但由于桂花的花期较短，大大缩短了桂花切花的观赏期，成为其发展切花的限制条件。目前，利用遗传工程在分子水平上控制花瓣衰老，延长花期的花卉育种已经得到了长足的发展。本实验建立了厚瓣金桂(O. fragrans ’Houban Jingui’)花瓣蛋白SDS-PAGE电泳实验体系，在蛋白质水平上对桂花切花衰老进行了研究。为桂花的遗传育种提供一些研究信息。 本试验研究结果如下： 1．蛋白电泳样品提取方法的确定。经过研究比较和改进，最后得到改良的丙酮沉淀法。利用该方法提取的蛋白样品得到的电泳结果杂质干扰少，蛋白带清晰且条带数较多。 2．桂花花瓣蛋白SDS-PAGE实验体系的建立。通过比较电泳结果调整分离胶浓度为11％，长度为10-11cm；浓缩胶浓度为3.9％，长度为1.5-2cm；样品上样量10μl，点样孔10×2.5×1mm~3；电泳时间2.5h左右，利用比较后选择的染色方法染色凝胶，得到了背景色浅、蛋白带清晰的凝胶图谱。在此基础上还对花瓣蛋白的双向电泳进行了初步试验。本试验还对单向SDS-PAGE结果图谱用2D软件包进行了分析，得到的蛋白质信息量比常规分析方法得到的要全面。 3．对切花花瓣衰老过程中蛋白的SDS-PAGE结果进行了分析，结果表明，39.8kD和27.5 kD多肽的表达水平从瓶插开始到结束蛋白条带基本保持稳定；37.6 kD多肽的表达水平随花瓣衰老而逐渐减弱：63.1kD、57.5 kD、51.5 kD多肽是随着花瓣的衰老而出现，且表达水平与花瓣的程度衰老成正比。对切花进行乙烯和ABA促衰处理导致51.5 kD和32.3 kD多肽的不同变化，说明这些多肽可能与它们引起衰老的不同机理有关。更多还原

【Abstract】 Osmanthus ( Osmanthus fragrans Lour.) is one of the traditional ornamental flower and is wildely used in gardens because of its specific fragrance .With the breed of large-flower varieties, osmanthus has become a kind of cut flower resource with good developing prospect. But osmanthus ornamental period is shortened by its short florescence, and this limits its development as cut flower. Nowadays, controlling the senescence of petal to prolong florescence by genetic engineering has obtained great development. This experiment established the SDS-PAGE experimental system for proteins of petal from osmanthus(O. fragrans ’Houban Jingui’), and studied the senescence of petal of osmanthus in protein level, which will bring some information for osmanthus genetic breed.The main results in this experiment were shown as follows:1. The determination of protein extraction method for electrophoresis. Based on studying and comparing other methods, an improved acetone-sediment method was achieved. By this method, the results of SDS-PAGE had clear protein bands with little background.2. Establishment of SDS-PAGE experimental system for petal proteins of osmanthus. After comparing the results of electrophoresis, the conditions of electrophoresis were modified : concentration of running gel was 11% with length of 9-10cm .the concentration of stacking gel was 3.9% with 1.5-2.0cm, the amount of added protein samples were 10ul in the vents of 10 X 2.5 X 1mm3, the running time was 2.5 hours or so. and gels were stained with the protocol selected from several staining methods. The proteins bands were clear and its background was low in the results of electrophoresis. Based on this, some preliminary study on 2-Dimensional electrophoresis for proteins of petal was carried out.3. The results of SDS-PAGE of proteins during senescent petal of cut osmanthus. The experiment analysis showed: 39. 8 kD and 27.5 kD peptides were stable from beginning to the end, 37.6 kD peptide decreased gradually and 63. 1kD, 57. 5 kD, 51.5 kD peptides appeared and increased with the senescence of petal. 51.5 kD, 32.3 kD paptides changed after ethylene and ABA treatment. This implies that there is some relationship between changes of peptides and senescence of petal.更多还原