芒果细胞超低温保存存活机理

李运合1,2, 李玉生3, 张金梅4, 陈晓玲4, 赵艳华3, 吴永杰3,*
1中国热带农业科学院南亚热带作物研究所, 广东湛江524091; 2农业部热带果树生物学重点实验室, 广东湛江524091; 3河北农林科学院昌黎果树研究所, 河北昌黎066600; 4中国农业科学院作物科学研究所, 北京100081

通信作者:吴永杰;E-mail: wuyongjie007@163.com

摘 要:

超低温保存是一个复杂的过程, 除了保存技术本身外, 一些因素如材料的选择及细胞生理状态的调控等也直接影响着保存后细胞的存活与再生。本文以芒果子叶诱导的胚性培养物和愈伤组织细胞为试材, 采用组织学观察、流式细胞分析和冷冻切片荧光观察等手段比较了其在超低温保存过程中的存活特性。结果表明胚性培养物细胞与愈伤组织细胞相比具有明显不同的细胞学特性, 从而导致其在超低温保存过程中具有很强的耐脱水和低温能力, 脱水处理30 min及超低温保存后存活率分别为96.7%和94.7%, 而愈伤组织存活率则为0。利用二乙酸荧光素(FDA)荧光染色法和冷冻切片技术, 观察到超低温保存后胚性培养物内存活细胞的分布。本研究结果证明试材细胞的生理状态对超低温保存的成功具有重要的影响, 对未来如何更准确地评价超低温保存的效果和确定最佳的超低温保存程序也具有重要意义。

关键词:芒果; 超低温保存; 愈伤组织; 胚性培养物

收稿:2016-07-11   修定:2016-09-05

资助:国家自然科学基金(31372053)。

The survival mechanism of mango cells during cryopreservation

LI Yun-He1,2, LI Yu-Sheng3, ZHANG Jin-Mei4, CHEN Xiao-Ling4, ZHAO Yan-Hua3, WU Yong-Jie3,*
1South Subtropical Crop Research Institute, Chinese Academy of Tropical Agricultural Sciences, Zhanjiang, Guangdong 524091, China; 2Key Laboratory of Tropical Fruit Biology, Ministry of Agriculture, Zhanjiang, Guangdong 524091, China; 3Changli Institute of Pomology, Hebei Academy of Agricultural and Forestry Sciences, Changli, Hebei 066600, China; 4Institute of Crop Science, Chinese Academy of Agricultural Sciences, Beijing 100081, China

Corresponding author: WU Yong-Jie; E-mail: wuyongjie007@163.com

Abstract:

In this study, two types of tissue cultures, callus and embryogenic cultures (EMs) were induced from cotyledon explants. The survival characteristics of both cultures during cryopreservation were investigated using histological observation, flow cytometric analysis, and FDA fluorescence dyeing and freezing section technology. Histological observation indicated that EMs had different cell characteristics compared with callus, which resulted in the stronger dehydration and cold resistance ability during cryopreservation. After 30 min dehydration treatment or further cryopreservation, the survival rates of EMs were 96.7% and 94.7%, respectively. By contrast, both survival rates of callus were 0. The distribution condition of survival cells of the EMs after cryopreservation was also observed by FDA fluorescence dyeing and freezing section technology. The results obtained here showed that the physiological statement of material cells was one essential factor for the success of cryopreservation. It would be helpful for the exactly evaluation of cryopreservation results and the optimization of cryopreservation procedures in the future study.

Key words: mango; cryopreservation; callus; embryogenic cultures

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