《植物生理学报》 2015, 51(8): 1322-1330

抗褐化剂对葡萄愈伤组织继代培养过程中酚类物质、相关酶及其基因表达的影响

饶慧云¹, 邵祖超¹, 柳海宁^1,*, 吴月燕¹, 刘蓉¹, 李学孚^1,2, 李美芹^1,2, 钱萍仙¹

¹浙江万里学院生物与环境学院, 浙江宁波315100; ²上海海洋大学水产与生命学院, 上海201306

通信作者：柳海宁；E-mail: nblhn@126.com；Tel: 13805863011

摘　要：

本文研究不同浓度的聚乙烯吡咯烷酮(PVP)、维生素C (Vc)、甘露醇、柠檬酸、硝酸银(AgNO₃)、活性炭(AC)和硫代硫酸钠(Na₂S₂O₃) 7种抗褐化剂对葡萄愈伤组织继代培养过程中抗褐化效果的影响。结果表明, 与对照比较, 除Na₂S₂O₃处理外, 其余处理的愈伤组织褐化率皆极显著下降。抗褐化的效果依次为PVP (2.0 g•L^-1)>甘露醇(20.0 g•L^-1)>AgNO₃ (0.02 g•L^-1)>Vc (0.006 g•L^-1)>AC (2.5 g•L^-1)>柠檬酸(0.6 g•L^-1), 其褐化率分别比对照降低62.28%、58.29%、53.13%、50.17%、42.33%和35.67%。葡萄愈伤组织的褐变与对羟基苯甲酸的含量和阿魏酸的含量呈正相关, 与咖啡酸的含量呈负相关。愈伤组织的褐化程度与多酚氧化酶(PPO)活性呈极显著负相关, 与苯丙氨酸解氨酶(PAL)活性呈极显著正相关; 与PPO基因的表达量呈极显著正相关, 与PAL基因的表达量呈显著负相关。因此, 在葡萄愈伤组织继代培养的过程中添加PVP (2.0 g•L^-1)、甘露醇(20.0 g•L^-1)和AgNO₃ (0.02 g•L^-1)等能够有效地防止愈伤褐变。

关键词：抗褐化剂; 葡萄; 愈伤组织; 酚类物质; 酶活性; 基因表达

收稿：2015-04-14   修定：2015-07-07

资助：宁波市重大科技专项(2015C110016)、浙江省科技厅项目(2013C24002)和宁波市科技创新团队项目(2011B82019)。

Effect of Browning Inhibitors on Callus Subculture of Phenolic Compounds, Enzyme and Gene Expression of Grape

RAO Hui-Yun¹, SHAO Zu-Chao¹, LIU Hai-Ning^1,*, WU Yue-Yan¹, LIU Rong¹, LI Xue-Fu^1,2, LI Mei-Qin^1,2, QIAN Ping-Xian¹

¹College of Biology and Environment, Zhejiang Wanli University, Ningbo, Zhejiang 315100, China; ²College of Fisheries and Life, Shanghai Ocean University, Shanghai 201306, China

Corresponding author: LIU Hai-Ning; E-mail: nblhn@126.com; Tel: 13805863011

Abstract:

Grape calli were selected as suitable materials and treated with different concentrations of a variety of different browning inhibitors, including polyvinyl pyrrolidone (PVP), vitamin C (Vc), mannitol, citric acid, silver nitrate (AgNO₃), activated carbon (AC) and sodium thiosulfate (Na₂S₂O₃), under grape callus subculture conditions to assess the effects of these agents on the browning rate of the calli. The results showed that the callus browning rate decreased significantly in all cases compared with the control, except for the callus subculture treated with Na₂S₂O₃. The anti-browning effects of the different browning inhibitors were as follow, PVP (2.0 g•L^-1)>mannitol (20 g•L^-1)>AgNO₃ (0.02 g•L^-1)>Vc (0.006 g•L^-1)>AC (2.5 g•L^-1)>citric acid (0.6 g•L^-1), and the associated browning rates had decreased by 62.28%, 58.29%, 53.13%, 50.17%, 42.33% and 35.67%, respectively. The para-hydroxybenzoic acid and ferulic acid contents were positively correlated with the browning rate of the callus, and negatively correlated with the content of caffeic acid. Negative and positive correlations were also found between the PPO, PAL activities and the browning rate of the callus, respectively. Furthermore, the expression levels of PPO and PAL were significantly positively and negatively correlated with the browning rate of the callus, respectively. These results therefore demonstrated that the addition of PVP (2.0 g•L^-1), mannitol (20 g•L^-1) or AgNO₃ (0.02 g•L^-1) could be useful to prevent callus browning during the subculturing of grape calli.

Key words: browning inhibitors; grape (Vitis vinifera); callus; phenolic compounds; enzyme activity; gene expression