植物抗寒冻基因工程研究进展

低湿寒害是限制农作物产量和分布的一种全球性的自然灾害。提高农作物的抗寒性具有重要意义。目前随着植物寒害机理、抗寒冻和冷驯化分子机理的深入发展,已研究发现了多种抗寒基因,包括各种抗寒调控基因和各种抗寒功能基因,从而使植物抗寒冻基因工程的研究与应用得以了广泛开展,以期最终有效地提高农作物的抗寒性,增加农业产量。本文综合概述了国内外有关植物抗寒冻基因工程的最新研究方向、进展及成就,并提出了此领域尚存在的一些问题及其前景展望。     

植物抗寒冻基因工程研究进展

低温寒害是限制农作物产量和分布的一种全球性的自然灾害,提高农作物的抗寒性具有重要意义。目前随着植物寒害机理、抗寒冻和冷驯化分子机理的深入发展,已研究发现了多种抗寒基因,包括各种抗寒调控基因和各种抗寒功能基因,从而使植物抗寒冻基因工程的研究与应用得到了广泛开展,以期最终有效地提高农作物的抗寒性,增加农业产量。本文综合概述了国内外有关植物抗寒冻基因工程的最新研究方向、进展及成就,并提出了此领域尚存在的一些问题及其前景展望。     

植物抗冻基因

介绍植物抗寒冻基因研究中一些已分离和鉴定出的低温诱导表达基因及其抗寒功能、低温信号转导以及调控方式的研究进展.     

植物抗寒冻的分子遗传与基因工程

植物抗寒冻的分子遗传与基因工程费云标,黄涛,舒念红,赵淑慧（中国科学院发育生物学研究所）植物抗寒冻的遗传学研究表明,小麦的２１对染色体中至少有１０对染色体关系到寒冻抗性,在染色体５Ａ和Ｄ,携带着影响小麦抗性的许多基因（Ｇｕｙ１９９０）。寒冻抗性的７０％的诱生能力涉及到染色体５Ａ（Ｓｉｎｇｈ等１９８８）。     

小麦原生质体在冰冻—化冻中的稳定性与其品种抗寒力的关系

对三个不同抗寒力小麦品种的分离原生质体进行了耐冻性的测定,结果指出,小麦原生质体在冰冻处理后的存活率与其品种的抗寒力成正相关。讨论了质膜的变化在寒害和抗寒机制上的作用,以及本结果作为鉴别植物抗寒力指标的意义。     

植物的低温蛋白

综述了与植物耐冻性有关的一些植物内源蛋白质或多肽 ,包括低温防护蛋白、抗冻蛋白、植物脱水素、膜关联耐冻性多肽蛋白质。结果表明 ,植物的耐冻性与其低温蛋白 (cold induced proteins)有着密切的关系 ,并指出了抗冻蛋白行使功能的两种可能的作用方式。同时 ,耐冻性与除低温外的其它环境胁迫因子的植物抗性如抗干旱、抗病虫、高盐耐性、乙烯耐性等密切相关     

植物抗寒性与抗寒基因的表达和调控

综合概述了国内外有关植物抗寒机理的研究动态,主要讨论了抗寒基因的表达与调控在植物抗寒性中的反应。此外,亦提出了有关植物抗寒机制研究领域值得深入研讨的问题。     

植物抗寒机理进展

本文综合概述阵内外有关植物抗寒机理研究的动态。主要讨论了植物抗寒性与细胞膜系、酶系多态笥及抗寒基因表达与调控之间的相关性。此外,亦提出了有关植物抗寒机制研究领域值得深入研讨的问题。     

植物抗寒机理研究进展

本文综合概述了国内外有关植物抗寒机理研究的动态,主要讨论了植物抗寒性与细胞膜系、酶系多态性及抗寒基因表达与调控之间的相关性。此外,亦提出了有关植物抗寒机制研究领域值得深入研讨的问题。     

植物抗寒机理研究进展

本文综合概述了国内外有关植物抗寒机理研究的动态,主要讨论了植物抗寒性与细胞膜系、酶系多态性及抗寒基因表达与调控之间的相关性。此外,亦提出了有关植物抗寒机制研究领域值得深入研讨的问题。     

Different responses to drought and freeze stress of Quercus ilex L. growing along a latitudinal gradient

The vulnerability to drought and freeze stress was measured in young plants of Quercus ilex L. growing in the field in two natural sites within the Italian distribution area of this species, i.e. Sicily (Southern Mediterranean Basin) and Venezia Giulia (Northeastern Italy), respectively. In particular, the resistance strategies adopted by Q. ilex to withstand the two stresses were estimated in terms of seasonal and/or diurnal changes in leaf conductance to water vapour (g_L), water potential (_L) and relative water content (RWC) as well as of xylem embolism in the stem and root hydraulic conductance (K_RL). Sicilian (SI) plants showed to reduce water loss by stomatal closure (g_L decreased) in summer, thus maintaining average RWCs at 88–90%. Moreover, SI plants showed considerable resistance to xylem cavitation in the stem (the loss of hydraulic conductance, PLC, was less than 12% throughout the year) and to maintain the hydraulic conductance of their roots (K_RL), constantly high even in summer. Plants growing in Venezia Giulia (VG plants), on the contrary, underwent leaf dehydration in the winter due to freeze stress so that RWC measured in April was still 78% on a diurnal basis. This was apparently due to consistent xylem embolism in the stem. In fact, PLC was as high as 40% between November and March. Only in the summer was PLC similar to that recorded in SI plants. Moreover, K_RL of VG plants decreased in November from about 1.5 to 0.8×10^–4 kg s^–1 m^–2 MPa^–1, i.e. about 50%, and in February K_RL dropped further to 0.4×10^–4 kg s^–1 m^–2 MPa^–1. On the basis of the above, we conclude that: (a) Q. ilex was more sensitive to freeze than to drought stress so that freeze stress can be considered as a factor limiting the distribution area of this species; (b) drought and freeze stress were faced by Q. ilex adopting two different resistance strategies, i.e. drought avoidance based on water saving in Sicily and freeze tolerance in Venezia Giulia.     

Freezing injury in potato leaves

Time-temperature profiles of freezing leaves from frost-resistant (Solanum acaule Bitt.) and frost-susceptible (Solanum tuberosum L. subsp. tuberosum Hawkes) types of potatoes did not reveal any major differences. The pattern of change in resistance of leaves to low voltage, low frequency current during freezing was different in the frost-resistant and susceptible leaves. In tissue sections from both types of leaves, cells freeze extracellularly at cooling velocities lower than 5 C per minute. Cells from leaves of resistant plants showed a higher osmotic pressure but not a higher water permeability than those from susceptible plants. The extent of injury caused by even very slow freezing was greater than that caused by equivalent isopiestic desiccation, particularly in susceptible leaves. The higher osmotic pressure in cells of leaves from resistant plants can account for the greater desiccation resistance but not for the frost resistance observed.     

Impact of freeze drying on drug resistance pattern of few Escherichia coli strains

Drug resistant strains of E. coli were freeze dried for long term preservation. Certain drug resistance markers were maintained after freeze drying while others were not. Streptomycin and sulphonamide resistance markers resisted freeze drying. Ampicillin, cephelaxin and neomycin resistances developed very frequently and except gentamicin all the markers were lost in varying percentage after freeze drying.     

Induction of Systemic Resistance to Tobacco Powdery Mildew by Tobacco Mosaic Virus, Tobacco Necrosis Virus or Ethephon

Local infections of either TMV or TNV in tobacco plants cv. Havana 425 (hypersensitive to TMV) proved effective in inducing systemic resistance to subsequent inoculation with the powdery mildew fungus Erysiphe cichoracearum DC. The proportion of leaf surface invaded by this pathogen and the amount of conidia it produced were both significantly lower in virus inoculated plants than in non-inoculated controls. However, the decrease in sporulation rate was less regularly observed than the reduction in leaf area infected. TMV was more effective than TNV in protecting tobacco plants from powdery mildew. E. cichoracearum is thus added to the list of challenge pathogens to which TMV or TNV are known to induce resistance in the host plants. Necrotic lesions caused to the leaves by local treatment with Ethephon (an ethylene-releasing compound) also conferred to tobacco some degree of systemic resistance to the same fungal pathogen, more frequently visible as a reduction of leaf area invaded. The protection due to the Ethephon lesions was in present experiments less marked than that of TMV. No effects against subsequent powdery mildew infection were obtained when point freeze necrotic lesions were provoked on the plants.     

Frost fatigue response to simulated frost drought using a centrifuge in Acer mono Maxim

In the coldest part of winter, water uptake is blocked by the frozen soil and frozen stems known as ‘frost drought’ causing severe embolisms in woody plants. Frost drought in stems was simulated in a centrifuge by a synergy between freeze–thaw cycles and the different tensions induced by changing the rotation speed. Frost fatigue was defined as a reduction of embolism resistance after a freeze–thaw cycle and determined from ‘vulnerability curves’, which showed percent losses of conductivity vs tension (positive value) or xylem pressure (negative value). Different tensions combined with a controlled freeze–thaw cycle were induced to investigate the effects on frost resistance over the course of year. During the growing season, Acer mono Maxim. developed significant frost fatigue, and a significant positive correlation was found between frost fatigue response and exogenous tension. During the dormant season, A. mono showed very high embolism resistance to frost drought, even under a tension of 2 MPa. When the exogenous tension was increased to 3 MPa while the stem was frozen, significant frost fatigue occurred. Longer freezing times had more serious effects on frost fatigue in A. mono. A hypothesis was raised that at the same low temperature, the severer the drought (higher tension) when stems were frozen, the higher frost fatigue response would be induced.     

Seasonality of cavitation and frost fatigue in Acer mono Maxim.

Although cavitation is common in plants, it is unknown whether the cavitation resistance of xylem is seasonally constant or variable. We tested the changes in cavitation resistance of Acer mono before and after a controlled cavitation–refilling and freeze–thaw cycles for a whole year. Cavitation resistance was determined from ‘vulnerability curves’ showing the percent loss of conductivity versus xylem tension. Cavitation fatigue was defined as a reduction of cavitation resistance following a cavitation–refilling cycle, whereas frost fatigue was caused by a freeze–thaw cycle. A. mono developed seasonal changes in native embolisms; values were relatively high during winter but relatively low and constant throughout the growing season. Cavitation fatigue occurred and changed seasonally during the 12‐month cycle; the greatest fatigue response occurred during summer and the weakest during winter, and the transitions occurred during spring and autumn. A. mono was highly resistant to frost damage during the relatively mild winter months; however, a quite different situation occurred during the growing season, as the seasonal trend of frost fatigue was strikingly similar to that of cavitation fatigue. Seasonality changes in cavitation resistance may be caused by seasonal changes in the mechanical properties of the pit membranes.     

Freeze/thaw stress in<Emphasis Type="Italic"> Ceanothus</Emphasis> of southern California chaparral

Freeze/thaw stress was examined in chaparral shrubs of the genus Ceanothus to determine the interactive effects of freezing and drought and to consider which is the more vulnerable component, the living leaves (symplast) or the non-living water transport system (apoplast). We hypothesized that where Ceanothus species co-occurred, the more inland species C. crassifolius would be more tolerant of low temperatures than the coastal species C. spinosus, both in terms of leaf survival (LT(50), or the temperature at which there is 50% loss of function or viability) and in terms of resistance to freezing-induced embolism (measurements of percent loss hydraulic conductivity due to embolism following freeze/thaw). Cooling experiments on 2 m long winter-acclimated shoots resulted in LT(50) values of about -10 degrees C for C. spinosus versus -18 degrees C for C. crassifolius. Freeze-thaw cycles resulted in no change in embolism when the plants were well hydrated (-0.7 to -2.0 MPa). However, when plants were dehydrated to -5.0 MPa, C. spinosus became 96% embolized with freeze/thaw, versus only 61% embolism for C. crassifolius. Stems of C. crassifolius became 90% and 97% embolized at -6.6 and -8.0 MPa, respectively, meaning that even in this species, stems could be more vulnerable than leaves under conditions of extreme water stress combined with freeze/thaw events. The dominance of C. crassifolius at colder sites and the restriction of C. spinosus to warmer sites are consistent with both the relative tolerance of their symplasts to low temperatures and the relative tolerance of their apoplasts to freeze events in combination with drought stress.     

Cold hardiness and water relations parameters in Rhododendron cv. Catawbiense Boursault subjected to drought episodes

We determined whether increase in cold hardiness of Rhododendron cv. Catawbiense Boursault induced by water stress was correlated with changes in tissue water relations. Water content of the growing medium was either maintained near field capacity for the duration of the study or plants were subjected to drought episodes at different times between 15 July and 19 February. Watering during a drought episode was delayed until soil water content decreased below 0.4 m³ m⁻³ then watering was resumed at a level to maintain soil water content between 0.3 and 0.4 m³ m⁻³. Cold hardiness was evaluated in the laboratory with freeze tolerance tests on detached leaves. Water relations parameters were determined using pressure-volume analysis. Exposure to drought episodes increased cold hardiness during the cold acclimation stage in late summer and fall but not during the winter. When water-stressed plants were re-watered to field capacity, the previous gain in cold hardiness gradually disappeared. Water relations parameters correlating with seasonal changes of cold hardiness included dry matter content (r =−0.67). apoplastic water content (r =−0.60), and water potential at the turgor loss point (r = 0.40). Changes of cold hardiness in water-stressed plants in reference to well-watered plants were correlated with changes of all water relations parameters, except for osmotic potential at full turgor (r = 0.13). It is proposed that water stress reduced the hydration of cell walls, thereby increasing their rigidity. Increased rigidity of cell walls could result in a development of greater negative turgor pressures at subfreezing temperatures and therefore increased resistance to freeze dehydration.     

Short term temperature drops do not enhance cold tolerance

To successfully transplant agricultural species in the spring, prior hardening is of great significance. Low, non-freezing temperature increases cold tolerance in many species. Also, diurnal temperature drops have been suggested to improve cold tolerance, as assessed by ultrastructural studies after short term freezing of leaf discs. Pre-treatment with lower day than night temperature prior to hardening has also been reported to enhance cold resistance in winter rape. This study investigated the effect of temperature drops on cold resistance of different species. In contrast to a period of continuous low temperature, short diurnal temperature drops did not enhance cold tolerance in Arabidopsis, swede, white cabbage or pea, compared to control plants. Exposure to low temperature of 6°C for 6 days increased cold tolerance by 2–5°C compared to plants exposed to diurnal temperature drops or control plants. Pre-treatment with diurnal temperature drops in the entire growth period prior to hardening with constant low temperature did not give any additional hardening in swede and pea. In conclusion, by freeze testing of whole plants under controlled conditions we have found no evidence supporting the hypothesis that diurnal temperature drops improve cold tolerance. However, temperature drops reduce plants size like shown earlier for a number of other species, and thus is a tool to produce compact, robust plants.     

不同小麦品种对低温胁迫的反应及抗冻性评价

以济麦19、济麦21、济南17等15个冬小麦品种为材料,对其在低温胁迫条件下功能叶和叶鞘超氧化物歧化酶(SOD)活性、过氧化物酶(POD)活性、过氧化氢酶(CAT)活性、丙二醛(MDA)含量与可溶性蛋白含量等生理指标进行测定,以功能叶各项指标的抗冻系数作为衡量抗冻性的指标,利用主成分分析、聚类分析对其抗冻性进行综合评价。低温胁迫条件下,不同冬小麦品种起身拔节期功能叶和叶鞘中SOD活性、POD活性和CAT活性均不同程度地上升,MDA含量和可溶性蛋白含量均下降。通过主成分分析和聚类分析,将15个冬小麦品种划分为3类:济麦19、山农8355属强抗冻类型;山农664、泰山9818、济麦21、济麦22、烟农24、烟农19、烟农21、汶农6号、鲁麦21、济南17属中度抗冻类型;其余3个品种(泰山23、聊麦18、临麦2号)属弱抗冻类型。