Abstract: The degenerate characteristics, including morphological, structural and vigorous changes of tobacco pollens were studied following different durations of cold treatment at 4℃ by using light and electronic microscopy, as well as TTC (triphenyltetrazolium chloride) assay. The following results were obtained: (1) After cold treatment, some of the pollens became empty and the cytoplasm of those pollens shrank. The longer the cold treatment lasted, the higher frequency of the degenerate pollens could be detected. (2) The degeneration of pollens was also detected at ultrastructural level. However, ultrastructural changes did not occurred synchronously with the duration of the treatment. Usually, dictyosomes and plastids degenerated first, followed by mitochondria and then the nucleus. During the process of the degeneration, vacuoles and endoplasmic reticulum (ER) played an important role. Vacuoles eroded and digested the surrounding cytoplasm gradually by partial or whole disappearance of tonoplasts. The proliferation of ER occurred as the time of the treatment increased. Simultaneously, the cisternae of ER became bulged, engulfing the cytoplasmic components. Finally, the cytoplasm shrank and converted into a dark disorganized mass when the pollen died. (3) Frequency of the vigorous pollens was detected by the TTC assay. It was shown that the frequency of the vigorous pollens decreased as the cold treatment went on. After 20 days of treatment, almost all the pollens lost their vigor. As a result, the degeneration of cold treated tobacco pollens occurred gradually and increasingly. It was PCD.

Key words: tobacoo, pollen, cold treatment, ultrastructure, programmed cell death

摘要： 利用环氧树脂半薄切片、超薄切片、电子显微镜技术以及氯化三苯四氮唑(TTC)法研究了烟草花粉经不同时间低温处理后的结构和活力的变化。在半薄切片中发现低温处理后一部分花粉细胞质收缩，整个细胞变瘪变空的衰退现象。随着处理时间的延长，衰退的花粉占花粉总量的比例也逐渐增大。超微结构观察表现出同样的但更加精细的结构变化趋势。从各种细胞器的变化来看，低温处理中首先是高尔基体、质体,然后线粒体发生衰退，细胞核可见的结构变化较晚。花粉衰退过程中液泡和内质网起重要作用。通过液泡膜的部分或完全消失，液泡不断侵蚀和消化邻近的细胞质。随着低温处理时间的延长，内质网大量增生、形成自噬泡，吞噬细胞质。最后花粉细胞质皱缩，变成一团深黑色无定形物质，花粉死亡。TTC法检测低温处理后花粉细胞活力变化的结果表明，随着低温处理进行，有活力的花粉占花粉总数的比例不断下降。到低温处理20d后，几乎所有的花粉都丧失了活力。结果表明：花粉在低温处理后的衰退过程是逐步的，有显著的时序性和特异性。这种衰退过程是细胞编程死亡。

关键词: 烟草, 花粉, 低温处理, 超微结构, 细胞编程死亡