Generation of reactive oxygen species during cryopreservation may improve Lilium x siberia pollen viability
Generation of reactive oxygen species during cryopreservation may improve Lilium x siberia pollen viability
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冷冻保存过程中活性氧的产生可能会提高百合 x 西伯利亚花粉的活力
DOI：
10.1007/s11627-014-9615-3
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发表时间：
2014
期刊：
影响因子：
--
通讯作者：
Shi Yin
中科院分区：
文献类型：
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作者：
Xu Jin;Liu Qian;Jia Mengxue;Liu Yan;Li Bingling;Shi Yin
在植物的生物和非生物胁迫过程中，活性氧（ROS）可能扮演两个非常不同的角色：高浓度的ROS可以加剧伤害，而低浓度的ROS可以激活防御反应。本研究的目的是探讨超低温保存后活性氧的产生与花粉生活力的关系。以2′，7 ′-二氯二氢荧光素二乙酸酯为荧光探针，采用流式细胞术检测了西伯利亚百合（Lilium×siberia）花粉中活性氧的产生。花粉生活力通过氯化2，3，5-三苯基四唑染色来测定。快速冷却使活性氧产生略有增加（新鲜花粉为26.13 ± 2.30），玻璃化冷冻使活性氧产生显著增加（49.74 ± 1.43;P< 0.01）。4.74vs.15.80快速冷却和玻璃化冷冻后的花粉生活力（分别为58.88 ± 3.76%和70.35 ± 2.90%）显著高于新鲜花粉（46.65 ± 1.61%;P <0.01）。在快速冷却之前，在不同温度下的冷驯化与ROS产生没有显着差异。然而，在4°C和−20°C下冷驯化相对于快速冷却而不驯化观察到存活率急剧下降（P< 0.01）。我们观察到在省略不同步骤的玻璃化冷冻处理中ROS产生无显著减少，而当省略卸载步骤时ROS产生显著减少（P< 0.05）。不进行装粉或脱水处理，花粉生活力极显著降低（P< 0.01）。在玻璃化过程中使用的不同溶液中加入200 U/ml过氧化氢酶时，在处理之间没有观察到显著差异的ROS产生或花粉生活力。综合分析表明，活性氧的产生与花粉生活力呈正相关（r= 0.651，P < 0.001）。因此，冷冻保存期间增加活性氧的产生可能会提高“西伯利亚”花粉的活力。
During biotic and abiotic stress in plants, reactive oxygen species (ROS) may play two very different roles: high ROS concentrations can exacerbate damage, whereas low concentrations can activate defense responses. The aim of this study was to investigate the relationship between ROS generation and pollen viability after cryopreservation. ROS generation was detected from ‘Siberia’ (Lilium×siberia) pollen using flow cytometry with 2′,7′-dichlorodihydrofluorescein diacetate as a fluorescent probe. Pollen viability was determined by 2,3,5-triphenyltetrazolium chloride staining. ROS generation was slightly increased by rapid cooling (26.13 ± 4.74vs.15.80 ± 2.30 for fresh pollen) and significantly increased by vitrification (49.74 ± 1.43;P< 0.01). Pollen viabilities after rapid cooling and vitrification were significantly increased (58.88 ± 3.76% and 70.35 ± 2.90%, respectively) over that of fresh pollen (46.65 ± 1.61%;P <0.01). No significant differences in ROS generation were associated with cold acclimation at different temperatures before rapid cooling. However, sharp decreases in viability were observed with cold acclimation at 4°C and −20°C relative to rapid cooling without acclimation (P< 0.01). We observed nonsignificant decreases in ROS generation among vitrification treatments that omitted different steps and a significant decrease when the unloading step was omitted (P< 0.05). Pollen viabilities were significantly reduced when the loading or dehydration steps were omitted (P< 0.01). No significant differences were observed in ROS generation or pollen viability among the treatments when 200 U/ml catalase was added to different solutions used in the vitrification process. Comprehensive analysis of all data indicated a positive correlation between ROS generation and pollen viability (r= 0.651,P< 0.001). Therefore, increasing ROS generation during cryopreservation may improve the viability of ‘Siberia’ pollen.