核桃属4 树种展叶期抗寒性鉴定

 以普通核桃（Juglans regia L.）、核桃楸（J. mandshurica Maxim.）、河北核桃（J. hopeiensis Hu）和黑核桃（J. nigra L.）4种核桃属植物的20份种质展叶期叶片为试验材料,采用电导法并配合Logistic方程确定其低温半致死温度（LT₅₀）;分析了LT₅₀与叶片解剖结构之间的相关性。结果表明,随着处理温度的降低,核桃叶片的相对电导率（REC）均呈上升趋势;供试核桃种质间的抗寒性差异较大,低温半致死温度在–8 ~ 2 ℃之间,在黑核桃、核桃楸及普通核桃中均有抗寒性强的种质资源。利用略高于大部分核桃种质LT₅₀的温度（0 ℃或–3 ℃）处理叶片,其相对电导率与低温半致死温度之间达到极显著正相关,而且相关系数高,此温度下处理12 h后叶片的相对电导率以及叶片的LT₅₀均可作为鉴定展叶期核桃种质抗寒性强弱的物理指标。核桃属不同种间的叶片结构类似,但各有特点。栅栏组织厚度和叶片总厚度与其LT₅₀之间均达到极显著负相关,可作为核桃叶片抗寒性鉴定的形态结构指标。     

苹果品种抗寒性测定方法的研究

采用电导法、恢复生长法、组织褐变法及Ｌｏｇｉｓｔｉｃ方程法对部分苹果品种进行了测定。提 出了以电解质渗出率 ４０％相对应的温度为枝条组织的临界致死温度（Ｃ． Ｌ． Ｌ．Ｔ）;通过组织褐变法 求得褐变温度（Ｂｒｏｗｎｉｎｇ Ｔｅｍｐｅｒａｔｕｒｅ,Ｂ．Ｔ．）;应用Ｌｏｇｉｓｔｉｃ方程求出拐点温度,即半致死温度（ＬＴ_(５０)）; 通过恢复生长法观察了各种处理温度下的萌芽率。综合以上测定结果,以ＩＢＰＧＲ分类标准为基础 将参试品种分为５级,列出了供试品种抗寒力的相对强弱顺序,并对抗寒性作了综合评价。     

山葡萄种质资源枝条的低温半致死温度研究

以山葡萄种质资源的成熟1年生休眠枝条为试材,测定了不同梯度低温处理下电导率的变化,采用相对电导率拟合Logistic方程,计算低温半致死温度(LT50),鉴定不同山葡萄种质资源的抗寒性。结果表明:用Logistic方程进行拟合后,拟合度高,所求低温半致死温度准确可信,可作为山葡萄抗寒性鉴定指标;试验的9份山葡萄种质资源间抗寒性存在一定差异,低温半致死温度范围为-31.98~-39.41℃,种质资源的抗寒性与枝条成熟度无明显相关性。     

黑龙江省5个主栽葡萄品种抗寒性鉴定

鉴定分析了黑龙江省5个葡萄品种1年生枝条解剖结构、电解质渗出率、可溶性糖含量等与葡萄抗寒性的关系。结果表明,供试葡萄品种抗寒性从强到弱为:贝达>京秀>京亚>U-尼昆>无核白;枝条电解质渗出率、可溶性糖含量、可溶性蛋白质含量、脂肪酸含量与抗寒性均呈正相关。     

四种园林树木抗寒性的比较分析

 以灯台树、复叶槭、流苏和山桐子幼树1年生休眠枝条为材料, 研究了不同冷冻处理条件下枝条的相对电导率和钾离子(K⁺ ) 相对渗出率, 利用Logistic方程计算了各树种的半致死温度(LT₅₀ ) , 并通过苗木生长恢复试验进行了验证。结果表明: 山桐子、复叶槭、流苏和灯台树4种枝条的相对电导率和K+相对渗出率均随处理温度的降低而呈“S”形上升, 其枝条的LT₅₀分别为- 11.5℃、- 24.9℃、- 20.7℃和- 29.8℃; 其抗寒性顺序依次为: 灯台树>复叶槭>流苏>山桐子, 与枝条恢复生长的试验结果基本一致。     

西府海棠抗寒性初步研究

本试验主要采用电导法,测定连续梯度降温处理的西府海棠一年生枝条的电导率,然后计算电解质渗出率,通过找出生物膜透性的变化规律来判断西府海棠枝条受冻害的程度,再配合徒手切片法观察组织结构变化和水培法观察生长情况来鉴定西府海棠枝条抗寒性的临界温度。     

德国鸢尾6个品种的耐寒性比较

 以6个不同花色的德国鸢尾（Iris germanica）品种为试材,通过田间鉴定与测定冬季自然降温过程中叶片相对电导率,结合Logistic方程计算各品种的半致死温度（LT₅₀）,评价其在自然降温过程中的耐寒性,并测定叶片中游离脯氨酸、丙二醛和可溶性蛋白的含量。结果表明：自然低温期6个品种寒害指数范围为1.20 ~ 3.65,半致死温度（LT₅₀）范围为﹣13.81 ~﹣5.73 ℃。耐寒性强的品种为‘Cherry Falls’和‘Bedtime Story’,其次为‘Brown Lasso’和‘Black Swan’,耐寒性弱的为‘China Dragon’和‘Caligula’。在自然降温过程中,6个德国鸢尾品种的低温半致死温度均随温度的下降而降低,下降幅度为1.46 ~ 6.51℃不等,并且叶片相对电导率随温度降低呈“S”型上升,丙二醛、脯氨酸和可溶性蛋白含量均呈现先上升后下降的趋势。     

电导法不同计量单位鉴定葡萄抗寒性研究

参照组织变褐法对电导法鉴定葡萄抗寒性所用不同计量单位(直接计量单位、电解质外渗率和统计计量单位)的可靠性和精确性进行了研究。结果表明,电解质外渗率计量单位比较可靠,其中伤害率(％)=处理电导率/处理煮沸电导率×100％的计量方式,适宜应用于大量材料的快速鉴定;伤害率(％)=(处理电导率/处理煮沸电导率-对照电导率/对照煮沸电导率)/(1-对照电导率/对照煮沸电导率))×100％计量方式所得结果精确可靠,宜于精细研究。     

苹果品种抗寒测定方法的研究

采用电志法、恢复生长法、组织褐变法及Ｌｋｏｇｉｓｔｉｃ方程法对部分苹果品种进行了测定。提出了以电解质渗出率４０％相对应的温度为枝条组织的临界致死温度（Ｃ．Ｌ．Ｌ．Ｔ）；通过组织褐变法求得褐变温度（Ｂｒｏｗｎｉｎｇ Ｔｅｍｐｅｒａｔｕｒｅ，Ｂ．Ｔ．）；应用Ｌｏｇｉｓｔｉｃ方程求出损点温度，即半致死温度（ＬＴ５０）；通过恢复生长法观察了各种处理温度下的萌芽率。综合以上测定，以ＩＢＰＧＲ分类标准为基础将     

持续低温胁迫下甜樱桃枝条电导率变化研究

用电导法测定了5个甜樱桃和1个酸樱桃的枝条细胞在经历持续低温胁迫的过程中电解质渗出率(即电导率)的变化,研究了5个甜樱桃品种的抗寒性。结果表明:所有樱桃品系的枝条细胞在接受持续-20℃低温胁迫处理过程中,细胞电解质渗出率先升高后降低,即细胞在接受临界致死温度附近的低温胁迫过程中,在一定的较短时间内细胞的自身恢复能力会快速提高。比较各品种之间电解渗出率的相对大小与分析细胞自身恢复能力情况得出,酸樱桃CAB、红艳和龙冠的抗寒力相对较强,是抗寒性较强品系、品种,红蜜、红灯和茉莉抗寒力相对较弱,是抗寒性较弱品种。     

菊花8个品种的低温半致死温度及其抗寒适应性

 通过测定秋冬季自然降温过程中菊花脚芽的叶片相对电导率（REC）,结合Logistic方程计算的半致死温度（LT₅₀）评价了8个不同花期菊花品种在不同降温时期的抗寒性,并通过生长恢复试验进行验证。结果表明,在自然降温过程中,8个菊花品种的低温半致死温度均随气温的下降而不断降低,但下降幅度因品种而异,从4.0～9.4℃不等。8个品种抗寒性由强到弱排序依次为'金陵黄鹤'>'银星'>'奥运晚霞'>'金陵之光'>'奥运锦云'>03（6）-16>03（6）-12>'奥运火炬',植株抗寒性与花期相关性不显著。11月底菊花脚芽恢复生长试验与当月半致死温度测定结果基本一致,表明半致死温度可作为菊花抗寒性评价的一个可靠指标。当温度降到-14℃时,供试品种的脚芽均不能恢复生长。     

低温对仁用杏雌蕊抗坏血酸-谷胱甘肽循环的影响低温对仁用杏雌蕊抗坏血酸-谷胱甘肽循环的影响

 利用人工智能模拟霜箱,设8 个温度（18、–1、–2、–3、–4、–5、–6、–7 ℃）,对 抗寒性不同的两个仁用杏（Prunus armeniaca Linn.）品种雌蕊的过氧化氢（H₂O₂）含量和抗坏血酸-谷胱 甘肽（AsA-GSH）循环进行了研究。结果表明,低温胁迫增加了仁用杏雌蕊H₂O₂ 含量,–7 ℃时抗晚霜 品种‘围选1 号’和晚霜敏感品种‘龙王帽’分别为对照（18 ℃）的 5.49 倍和7.22 倍。低温期间,‘围 选1 号’雌蕊AsA 等酶含量及APX 等酶活性均高于‘龙王帽’,–7 ℃时,‘围选1 号’雌蕊AsA、MDHA、 DHA、GSH 和GSSG 含量比对照分别下降了66%、69%、48%、52%、53%,‘龙王帽’下降了78%、79%、 59%、66%、65%,–7 ℃时,‘围选1 号’雌蕊APX、MDHAR、DHAR、GPX、GR 活性、AsA/DHA、 GSH/GSSG 分别是对照的2.87 倍、1.07 倍、1.42 倍、1.60 倍、1.20 倍、65%和1.03 倍,‘龙王帽’是对照 2.41 倍、96%、1.13 倍、1.08 倍、98%、54%、97%。结果表明‘围选1 号’雌蕊在低温期间有较高的AsA-GSH 循环效率,可有效抑制H₂O₂ 的积累,这可能是其抗晚霜能力较强的重要原因之一。     

Protocol: Streamline cloning of genes into binary vectors in Agrobacterium via the Gateway® TOPO vector system

Background

Freezing tolerance is an important factor in the geographical distribution of plants and strongly influences crop yield. Many plants increase their freezing tolerance during exposure to low, nonfreezing temperatures in a process termed cold acclimation. There is considerable natural variation in the cold acclimation capacity of Arabidopsis that has been used to study the molecular basis of this trait. Accurate methods for the quantitation of freezing damage in leaves that include spatial information about the distribution of damage and the possibility to screen large populations of plants are necessary, but currently not available. In addition, currently used standard methods such as electrolyte leakage assays are very laborious and therefore not easily applicable for large-scale screening purposes.

Results

We have performed freezing experiments with the Arabidopsis accessions C24 and Tenela, which differ strongly in their freezing tolerance, both before and after cold acclimation. Freezing tolerance of detached leaves was investigated using the well established electrolyte leakage assay as a reference. Chlorophyll fluorescence imaging was used as an alternative method that provides spatial resolution of freezing damage over the leaf area. With both methods, LT₅₀ values (i.e. temperature where 50% damage occurred) could be derived as quantitative measures of leaf freezing tolerance. Both methods revealed the expected differences between acclimated and nonacclimated plants and between the two accessions and LT₅₀ values were tightly correlated. However, electrolyte leakage assays consistently yielded higher LT₅₀ values than chlorophyll fluorescence imaging. This was to a large part due to the incubation of leaves for electrolyte leakage measurements in distilled water, which apparently led to secondary damage, while this pre-incubation was not necessary for the chlorophyll fluorescence measurements.

Conclusion

Chlorophyll fluorescence imaging is an alternative method to accurately determine the freezing tolerance of leaves. It is quick and inexpensive and the system could potentially be used for large scale screening, allowing new approaches to elucidate the molecular basis of plant freezing tolerance.     

河北核桃( Juglans hopeiensis Hu)光合特性的研究

 以河北核桃‘艺核1号’等为试材, 利用CI-301PS便携式光合作用测定仪, 在田间条件下,研究了河北核桃的光合特性。结果表明: ‘艺核1号’光饱合点为1643 μmol·m^{- 2} ·s^{- 1}左右, 光补偿点为50 μmol·m ^{- 2} ·s^{- 1}左右, 最适温度为29℃; 光合速率日变化为“中午降低型”的双峰曲线; 净光合速率的季节变化为双峰曲线, 第1峰出现在5月中旬, 第2峰出现在8月上旬; 影响光合速率日变化的主要因子依次是气孔导度、光合有效辐射、叶温、胞间CO₂ 浓度和蒸腾速率。河北核桃净光合速率低于普通核桃, 接近于核桃楸。     

Comparative analysis of laboratory freezing methods to establish cold tolerance of detached rhizomes and intact crowns in garden chrysanthemums (Dendranthema × grandiflora Tzvelv.)

Since 1926, the University of Minnesota herbaceous perennial breeding program has released N = 84 garden chrysanthemum cultivars (Dendranthema × grandiflora) with important traits for northern temperate climates, such as winter hardiness. Recent breeding objectives have identified the need for development of non-destructive phenotypic markers and destructive laboratory freezing tests for co-selection of cold tolerance in Dendranthema, Gaura, and other herbaceous perennial flower crops. Such methods have become critical to flower breeding programs in northern temperate regions during periods of above-average winter temperatures and minimal snow cover due to the ‘el Niño’ effect. Two different, destructive laboratory freezing tests were evaluated for their effectiveness in determining cold tolerance. Acclimated crowns of n = 6 garden chrysanthemum genotypes, ranging from hardy to non-hardy in USDA Z3-4, were used in Omega Block (using detached, emergent rhizomes) and chamber (using entire, intact crowns with emergent, non-emergent rhizomes) freezing test methods. Comparative winter survival in the field was monitored over locations and years. Cold tolerance was assessed at 0 to −12 °C with varying ramp and soak time periods. LT₅₀ temperatures and number of living emergent rhizomes were also determined. Rhizome quality at 1, 3, and 5 cm depths was rated for regrowth on a 0 (dead) to 5 (undamaged) scale. The chamber freezing method was the most powerful to discern accurate LT₅₀ values. Cold tolerant genotypes included ‘Duluth’ and Mn. Selection 98-89-7 (LT₅₀ = −12 °C). Four genotypes were rated as non-hardy (LT₅₀ = ≤−10 °C). Cold-tolerant genotypes also had significantly higher regrowth ratings for rhizomes at 1 and 3 cm depths. Future research will implement the chamber freezing method to assay the inheritance of winter hardiness in intact crowns of segregating populations.     

低温胁迫下佛手半致死温度测定和抗寒性分析

 以‘青皮’和‘矮化’佛手(Citrus medica var. sarcodactylis Swingle)为试材,测定不同低温下处理24 h后,叶片的电解质外渗率(REC),计算半致死温度(LT₅₀);在半致死温度下设置不同处理时间,测定REC、超氧化歧化酶(SOD)活性、游离脯氨酸(Pro)和丙二醛(MDA)含量,进行抗寒性分析。结果表明：佛手REC随温度降低及处理时间延长呈“S”形上升, LT₅₀在-4~ -5℃之间;在LT₅₀下,随着处理时间的延长SOD活性呈先上升后下降的趋势, Pro和MDA含量逐渐上升;12~ 24 h为佛手在LT₅₀下冷害发生的临界时间。     

The fractal spectrum of leaf colour as a tool for measuring frost hardiness in plants

Summary

An image analysis method, based on the calculation of the fractal spectrum of leaf colour, was evaluated for its potential use in assessing frost hardiness in plants. The technique developed consisted of a) digitization of the leaves after freezing treatments, b) splitting of the images obtained into the three components of the RGB (red, green and blue) colour system and c) calculation of the fractal spectrum for each colour component of the leaf. It was then tested to assess the freezing hardiness of acclimated and non-acclimated Callistemon linearis plants. The analysis system consisted of a scanning device, a personal computer and the image analysis algorithm. All the fractal parameters showed a sigmoidal relationship with temperature. From the inflection point, the LT₅₀ calculated for acclimated and non-acclimated plants, was –7.5 and –4.8°C, respectively. The LT₅₀ calculated from the electrolyte leakage test on the same material was about 2°C warmer than the LT₅₀ obtained from fractal analysis. The role of fractal analysis of leaf colour as an additional, rapid and inexpensive method for identifying freezing damage is discussed.     

苹果砧木枝条电阻抗参数与其抗寒性的关系

 以18株2年生苹果砧木西府海棠（Malus micromalus Makino）实生后代单株为试材,研究了不经冷冻处理的枝条的电阻抗（EIS）参数与其抗寒性（电导法测定枝条冷冻半致死温度）的关系。结果表明,3个时期（3月、7月、11月）枝条的EIS参数高频电阻率r和低频电阻率r1、胞外电阻率re、弛豫时间τ与抗寒性呈显著负相关（r =﹣0.737** ~﹣0.877**）,在11月时胞内电阻率ri与抗寒性也呈显著负相关（r =﹣0.857**）,表明在非冷冻条件下可采用枝条的电阻率r和r1、胞外电阻率re、弛豫时间τ估测苹果砧木实生后代枝条的抗寒性。通过比较3个时期各参数的最终通径系数,得出re为估测枝条抗寒性最佳的参数。