Culture of Plant Protoplasts for Studies of Senescence

植物衰老的最明显迹象是花、果实、叶和根的枯萎和最终死亡。衰老是一个复杂的过程，涉及环境、生理和遗传因素，其机制尚不完全清楚。在这个项目中，将开发一种独特的模型来研究植物衰老，最终目的是阐明其分子机制。植物衰老对植物的整体经济、果实和谷物成熟、作物生产和作物的收获后贮藏都很重要。在叶片衰老过程中，蓝烟叶肉细胞中的叶绿体失去功能，而气孔保卫细胞中的叶绿体不丧失功能。在这个项目中，叶肉细胞和保卫细胞将以保存分化细胞类型的独特属性的方式进行培养。为了优化细胞存活和保存每种细胞类型特有的生理特性，将测试各种营养、渗透、pH、光和细胞密度条件。一旦建立了培养，将确定融合两种细胞类型的原生质体和培养融合杂交种的条件。将用显微荧光分光光度法监测融合杂种保卫细胞叶绿体的功能，以确定衰老的叶肉细胞是否含有能够启动保卫细胞叶绿体衰老的因子。

The most obvious signs of plant senescence are the withering and eventual death of flowers, fruits, leaves and roots. Senescence is regulated by complex processes involving environmental, physiological and genetic factors whose mechanisms are incompletely known. In this project, a unique model will be developed for the study of plant senescence, with the eventual goal of elucidating the molecular mechanisms. Plant senescence is important to the overall economy of the plant, fruit and grain ripening, crop production, and post-harvest storage of crops. Chloroplasts in mesophyll cells of Nicotiana glauca lose functionality during leaf senescence, but those in stomatal guard cells do not. In this project, mesophyll and guard cells will be cultured in ways that preserve the unique properties of the differentiated cell types. To optimize cell survival and preservation of physiological properties unique to each cell type, a variety of nutrient, osmotic, pH, light, and cell density conditions will be tested. Once cultures are established, conditions will be identified for fusing protoplasts from the two cell types and for culturing fusion hybrids. The functionality of guard cell chloroplasts in fusion hybrids will be monitored by microfluorospectrophotometry to determine whether senescing mesophyll cells contain factors capable of initiating guard cell chloroplast senescence.***//