马铃薯茎尖超低温保存中细胞死亡模式及其调控研究

Cryopreserved explants often exhibit lower recovery rates which significantly hampers wide application of plant cryopreservation. The cryo-associated injuries are responsible for the cell death. It is not clear that only necrosis or programmed cell death is also involved during cryopreservation induced cell death. The effect mechanisms of cryo-associated injuries on modes of cell death have also not yet revealed. The aims of the present study are to identification and analysis the two modes of cell death by investigating the morphology and DNA cleavage characteristics in cells of potato shoot tips, to analyze and compare the cell injuries suffered from ice crystals, cryoprotectant and ROS when applying two cryogenic approaches, thus understanding the effect of cryo-associated injuries on modes of cell death, in addition to identify the major injury and mediated factor causing different death modes. Furthermore, the modulation mechanism could be revealed by regulating the major injury and mediated factors, which cause cell necrosis and induce programmed cell death, respectively. Thus the modulation means of different modes of cell death can be developed. The results are valuable in providing newly ideas and strategies for overcoming the bottleneck problems related to higher rates of cryopreservation failure.

植物材料液氮冷冻后存活率低是限制超低温保存广谱应用的瓶颈问题，超低温保存伤害是造成冻存材料死亡的根本原因，但其细胞究竟是仅发生坏死还是兼有程序性死亡，目前并不清楚，超低温保存伤害的不同类型及其程度对细胞死亡模式的影响机制也尚未阐明。本项目拟以马铃薯为材料，通过研究超低温保存中死亡细胞的组织学和超微结构特征及DNA降解模式，对茎尖细胞的死亡模式进行鉴定和分析；应用透射电镜、差示扫描量热和组织化学定位技术解析和比较不同超低温保存方法中冷冻保护剂、冰晶和ROS三种关键因素对茎尖细胞造成的伤害，通过分析不同伤害因素对细胞死亡模式的影响，揭示细胞坏死的主导伤害和程序性死亡的介导因素；进一步通过研究主导伤害和介导因素的调节方法及对两种死亡模式的影响，建立超低温保存中细胞不同死亡模式的调控方法。研究可为解决超低温保存材料存活率低的瓶颈问题提供新思路和新策略。