TTC染色鉴定葡萄抗寒性方法的优化与应用  

Optimization of TTC Staining of Grape Branch and Its Application in the Evaluation of Cold Resistance

作　　者：王涵雅 李欣 毛娟[1] 马宗桓[1] 韦霞霞 陈佰鸿[1] WANG Hanya;LI Xin;MAO Juan;MA Zonghuan;WEI Xiaxia;CHEN Baihong(College of Horticulture,Gansu Agricultural University,Lanzhou 730070,China)

机构地区：[1]甘肃农业大学园艺学院,兰州730070

出　　处：《西北植物学报》2022年第10期1760-1768,共9页Acta Botanica Boreali-Occidentalia Sinica

基　　金：甘肃省科技重大专项(18ZD2NA006)。

摘　　要：以‘左山1号’、101-14、3309C、SO4、140R和‘黑比诺’等葡萄砧木和品种直径为0.5~0.7 cm的一年生枝条为材料,经低温梯度(0℃、-14℃、-19℃、-24℃、-29℃和-34℃)处理后,对TTC染色的温度和时间进行优化并观察统计葡萄枝条不同组织的存活情况,测定枝条的相对电导率,以及韧皮部和木质部的可溶性蛋白、游离脯氨酸、可溶性糖、淀粉含量和束缚水自由水比例(束自比)5个生理指标,分别用枝条纵切面染色面积、相对电导率拟合Logistic方程计算枝条的半致死温度(LT_(50))来评价枝条的抗寒性,同时通过隶属函数法综合评价枝条韧皮部和木质部抗寒性,并将3种方法的评价结果进行比较,以建立一种直观高效鉴定葡萄品种抗寒性的方法。结果表明:(1)依据低温胁迫下各品种葡萄砧木扦插枝条的萌芽率和生根率表现,其抗寒性由强到弱依次为‘左山1号’、101-14、3309C、SO4、140R和‘黑比诺’。(2)葡萄枝条TTC活力染色的最佳条件为pH=7.0,0.5%TTC-0.1 mol/L磷酸缓冲液在35℃下避光染色36 h。随着胁迫温度的降低,各品种枝条纵切面染色面积均逐渐降低,根据优化TTC法获得‘左山1号’、101-14、3309C、SO4、140R和‘黑比诺’枝条的LT_(50)分别为-31.38℃、-26.51℃、-26.10℃、-23.60℃、-23.33℃和-19.26℃。(3)随着胁迫温度的降低,各品种枝条的相对电导率逐渐增加,且‘黑比诺’的相对电导率基本保持最高、增幅最大(51.93%),而‘左山1号’的相对电导率始终最低、增幅最小(44.07%);根据相对电导率获得‘左山1号’、101-14、3309C、SO4、140R和‘黑比诺’枝条的LT_(50)分别为-30.02℃、-26.40℃、-25.75℃、-23.16℃、-21.13℃和-17.72℃。(4)通过5种生理指标进行枝条抗寒性隶属函数综合性评价结果显示,同一品种中韧皮部抗寒性强于木质部,同一部位不同品种的抗寒性表现为:‘左山1号’>101-14>3309C>SO4>140R>‘黑比诺Annual branches with a diameter of 0.5-0.7 cm from grape rootstocks and varieties of ’Zuoshan No.1’, 101-14, 3309 C, SO4, 140R, ’Pinot noir’, etc. were used as the materials. After the treatment of low temperature gradient(0 ℃,-14 ℃,-19 ℃,-24 ℃,-29 ℃ and-34 ℃), the temperature and time of TTC staining were optimized, and the survival of different tissues from grape branches was observed and counted. The relative electrical conductivity of the branches, the contents of soluble protein, free proline, soluble sugar and starch in phloem and xylem, and the ratio of bound water to free water(bundle to body) were measured. The semi-lethal temperature(LT) of branches was calculated by Logistic equation fitting the dyeing area of longitudinal section and relative electrical conductivity to evaluate the cold resistance of branches. Meanwhile, the cold resistance of the phloem and xylem of the branches was comprehensively evaluated by the physiological index membership function method. The evaluation results of the three methods were compared to establish an intuitive method for identifying the cold resistance of grape varieties. The results showed that:(1) the optimal conditions for TTC viability staining of grape shoots were pH=7.0, 0.5% TTC-0.1 mol/L phosphate buffer at 35 ℃ for 36 h in the dark. With the decrease of stress temperature, the dyeing area of the longitudinal section of each variety decreased gradually. According to the optimized TTC method, the LTof branches form ’Zuoshan No 1’, 101-14, 3309 C, SO4, 140 R, ’Pinot noir’ were-31.38 ℃,-26.51 ℃,-26.10 ℃,-23.60 ℃,-23.33 ℃ and-19.26 ℃, respectively.(2) With the decrease of stress temperature, the relative conductivity of branches of each variety gradually increased, and the relative conductivity of ’Pinot noir’ basically maintained the highest and the largest increase(51.93%), while the relative electrical conductivity of ’Zuoshan No.1’ always had the lowest and the smallest increase(44.07%). According to the rela

关 键 词：葡萄 抗寒性 TTC染色 半致死温度 韧皮部 木质部 综合评价 

分 类 号：Q256[生物学—细胞生物学] Q945.78[农业科学—果树学] S663.1[农业科学—园艺学]