种子贮藏中细胞死亡机制研究
陈虹颖
学位类型博士
导师李唯奇
2010-05
学位授予单位中国科学院研究生院
学位专业植物学
摘要种子作为承载植物生命的载体,并可以在有限的空间里最大限度地保存物种的遗传多样性,开展植物种子的收集保存已经成为当今植物多样性保护的一个重要手段。论文运用生理、生化和分子研究手段,对正常型种子老化过程和非正常型种子(即中间型和顽拗型)的干燥过程所导致的种子活力丧失,最终死亡,两个种子生物学研究领域的热点和难点问题进行了探索。结果发现:第一,“正常型”豌豆种子老化过程属于程序性死亡过程。本研究首次运用基因芯片技术,对种子老化过程进行了转录水平的剖析研究。为深入了解种子老化的机制奠定了基础。此外,谷光甘肽(glutahione)在“正常型”种子老化过程中扮演重要作用。研究检测了豌豆种子老化全过程中,谷光甘肽氧化还原状态的变化,结果显示,当种子活力开始下降时,谷光甘肽半细胞氧化还原电势(EGSSG/2GSH)明显上升, 表明在老化过程中氧胁迫累积,谷光甘肽是老化过程中种子活力的指示剂。说明谷光甘肽氧化还原系统在种子老化过程中,有举足轻重的作用。第二,活性氧可以缓解干燥导致的萌发率降低。论文以储藏特性为中间型的茶(Camellia sinensis var. assamica)种子为研究对象,在不同干燥程度的种子中加入外源ROS后发现:ROS可以减缓干燥对种子造成的伤害。第三,正常和顽拗型种子在脱水中分别经历可逆和不可逆的膜伤害。细胞膜对环境敏感,调整膜脂分子组成是细胞应对环境影响的主要方式。一个普遍的规律是,结构特殊的磷脂酸(phosphatidic acid,PA)分子对温度和水分十分敏感,其含量升高说明细胞膜受到了伤害。以贮藏行为不同的豌豆(Pisum sativum,正常型)、茶(Camellia sinensis var. assamica,中间型)和板栗(Castanea mollissima,顽拗型)为代表,利用脂类组学方法及聚类分析等手段,比较分析其变化规律与贮藏行为的关系。正常型和顽拗型胚轴的PA对脱水的响应模式不同。证据提示,正常型种子的萌发率不受脱水的影响,可能是脱水导致的细胞膜伤害是可逆的;顽拗型种子的萌发率在脱水中急剧下降,可能是其中脱水诱导的细胞伤害是不可逆的,最终导致了细胞死亡。暗示PA有可能成为种子干燥过程中的一个活力指示剂。
资助项目Seeds contain the entire genetic information of plants, which enable to develop the technology of seed preservation for the conservation of plant diversity. The mechanism of cell death in seeds, which are induced by ageing of orthodox seeds and desiccation of recalcitrant and intermediate seeds are two important areas in seed biology but poorly understood.Those two areas were investigated in this dissertation, and Pisum sativum, Camellia sinensis var. assamica, Castanea mollissima were chosen to represent orthodox, intermediate and recalcitrant seeds respectively as studied models. Biophysical, biochemical and molecular methods were employed to investigate the cell death mechanism during seeds ageing and desiccation.First of all, Ageing induced porgrammed cell death in seeds.The morphology change, nucleic acid integrity, and gene expression during ageing of Pisum sativum seeds were investigated.The data above confirmed that PCD is an underlying mechanism of seed mortality. Moreover, glutathione played an important function during seeds ageing.An increase in glutathione redox potential (EGSSG/2GSH) while seed lost viability was discovered during ageing of P. sativum seeds. Seed viability decreased 50% when EGSSG/2GSH attained values of -180 to -160 mV, which proved that EGSSG/2GSH was a marker for seeds viability. Secondely,Reactive oxygen species plays different roles in the seed germination after desiccation. After different desiccation treatments on the tea (C. sinensis var. assamica) seeds, the relationship between ROS dose and the viability of seeds after desication treatment was investigated. The results demonstrated that the presence of ROS could released the damage of desiccation on the tea seeds. Thridly,the composition of membrane lipid species and the change during desiccation in seeds with different storage behavior. Cell membrane is very sensitive to environment, and adjust the composition of lipid molecules is one of the main methord for cell to response the environmental changes. There is a rule that phosphatidic acid (PA), with special structure, is very sensitive to temperature and water stress, and the increase of PA content indicates the damage of the cell membrane. ESI/MS-MS technology was used to analysis and compare the composition and changes membrane lipid species during desiccation treatments of Pisum sativum, Camellia sinensis var. assamica, Castanea mollissima seeds which represented orthodox, intermediate and recalcitrant seeds respectively. The results showed that the responses of PA to dehydration in embryonic axis of orhtordox and recalcitrant seeds were different. The percentage increasement of PA in orthodox seed during desiccation was not incredible and the level of PA decreased and achieved the same level as controled, while PA level increased dramatically without showing any decrease trend in recalcitrant seed, which indicated that the damage to membrane of recalcitrant seeds was inevitable. Those results indicted that the dehydration might induced reversible memebrane damage in the orthodox seed, which may result in no affect on the seed viability, while it might induced irreversible membrane damage to the recalcitrant seed, which inducned the dramatically reduce of viabiligy and cell death in the end. It was the first time to investigate and analysis systematically the composition and changes of membrane lipids of different storage behavior seeds during desiccation. It was first time to report the relationship between PA and the seeds storeage and the change of PA level in seeds could be used as the marker to diagnose.the property of seeds for desiccation sensitivity or desiccation tolerance
语种中文
文献类型学位论文
条目标识符http://ir.kib.ac.cn/handle/151853/16096
专题昆明植物所硕博研究生毕业学位论文
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陈虹颖. 种子贮藏中细胞死亡机制研究[D]. 中国科学院研究生院,2010.
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