外源NO对镉胁迫下紫花苜蓿种子萌发及生长的影响
发布时间：2018-09-19 13:36
【摘要】：镉(Cd)对于动物和植物组织来说是一种有毒的和非必需的重金属。对植物在应对重金属伤害及植物对重金属的抗性机理的探索与研究是非常重要的课题,只有彻底了解植物在应对重金属胁迫伤害及其对重金属的抗性机理,才可能提出可以让植物避免遭受重金属伤害的有效的措施和提高植物抗重金属能力的重要方法,这样才有益于治理土壤中的重金属污染,保护地球生态环境和生产绿色健康的食品。外源一氧化氮(NO)是一种信号分子,对调节非生物和生物胁迫下各种生理反应具有重要作用。导致这些生理反应的重金属胁迫的信号在植物体内如何传递,NO在其中起什么作用,怎样调动防御系统并最终影响植物生长发育、生理代谢和基因表达等过程的,如何通过人为调控植物对重金属的耐受机制整体过程中的一些过程,来提高植物对重金属的耐性,增加或减少对重金属的吸收,并应用于重金属污染区的植物修复和农业生产?目前未见研究报道。紫花苜蓿(Medicago sativa L.)作为多年生的一种草本植物以其豆科紫花苜蓿属独有的特点在牧草业被认为是牧草之最。广泛分布在世界温带地区,27℃的温度最适于幼苗生长。其广泛的根系使其具有耐旱性,可以增强降解细菌的活性,促进多氯联苯(PCBs),多环芳烃(PAH)和三硝基甲苯(TNT)等几种有机污染物的生物降解率,尤其是在含有较低含量有机质的土壤中。紫花苜蓿含有丰富的维生素A、C、E、K及矿物质和微量元素,生长快速,根系发达、寿命长以及抗性好等特性,是有毒金属植物修复的有前景的植物。已有研究表明,紫花苜蓿对重金属的抗性比较强并且具有很好的吸收富集能力,其中对Cd的吸收系数最大,已有研究证明紫花苜蓿对Cd污染土壤的修复能力最好,被认为是重金属污染植物修复的环保技术之一。为探讨硝普钠(SNP,外源NO供体)对Cd在紫花苜蓿幼苗萌发过程中的毒性影响及其在植物修复Cd污染土壤的应用,本研究测定了氧化应激水平、抗氧化酶活性、水解酶活性、贮藏物质含量、鲜重、干重量和Cd的吸收等参数,分析了外源NO对Cd胁迫下紫花苜蓿种子萌发及生长的影响。萌发和生长实验进行5天,每隔24小时记录并测定一次各个指标。为揭示外源NO可以显著缓解Cd胁迫下紫花苜蓿萌发期间Cd毒性提供了证据。用不同浓度的CdCl_2对种子和幼苗进行预处理,结果表明CdCl_2对紫花苜蓿种子的发芽和幼苗生长的毒性作用随着其浓度的增加而增加。30μM的CdCl_2对紫花苜蓿的幼苗生长有明显的抑制作用而不会对其发芽率造成明显的影响。高浓度的CdCl_2会导致发芽率严重下降,从而减少了每组实验中所产生幼苗的可用样本。因此本研究采用30μM CdCl_2模拟Cd胁迫。在30μM CdCl_2模拟的Cd胁迫下,以25、50、100、200、300、400和500μM SNP分别处理紫花苜蓿种子5天,每隔24小时记录并测定一次以下指标:发芽率、发芽指数、壮苗指数、根长和芽长作为萌发参数;脯氨酸(Pro)水平,相对电导率(REC),脂质过氧化(MDA)和活性氧(ROS)如H2O2和O2-产生速率作为Cd作用下的氧化应激反应参数;超氧化物歧化酶(SOD)、过氧化物酶(POD)、抗坏血酸过氧化物酶(APX)、过氧化氢酶(CAT)和谷胱甘肽还原酶(GR)等抗氧化酶活性作为抗氧化机制的参数;α-淀粉酶和β-淀粉酶活性作为水解酶的参数;可溶性蛋白、可溶性淀粉、可溶性糖和还原性糖作为储存物质的参数。1、外源NO能够缓解Cd胁迫对紫花苜蓿种子萌发和幼苗生长的抑制作用。30μM Cd胁迫加速了储存物质的分解,显著降低了种子发芽指数和幼苗活力指数,抑制了幼苗生长,外源施用的不同浓度的SNP表现出显著的缓解重金属胁迫的作用,具有缓解胁迫和促进植物生长的能力。在最佳浓度下,缓解作用呈显著的线性增加,而超过最佳浓度时则使植物生长受抑制。这种反应更可能是通过有效的外源NO激活植物防御或耐受机制的结果。这证实了以前关于砷胁迫下的种子发芽过程中的NO的保护作用的研究。对Cd胁迫条件下水稻种子萌发及其幼苗生长的研究已经阐明了外源NO对Cd毒性的重要性,SNP能够显著提高发芽率,发芽指数和幼苗活力指数,在芝麻苗中也已经报道了类似的结果。在本研究中,外源性NO成功缓解Cd损伤,可能也以与其他生物或非生物胁迫类似的方式运行。然而,在SNP的最佳浓度之上,植物或幼苗表现出相同的防御机制,这类似于多年生黑麦草对Cd胁迫的防御机制。200μM SNP能有效缓解30μM的Cd对紫花苜蓿种子萌发和有生长的抑制作用。2、外源NO通过激活抗氧化系统,清除活性氧,缓解Cd胁迫下紫花苜蓿的氧化损伤。Cd能够诱导幼苗中产生ROS,如O2-和H2O2,它们导致膜脂过氧化,电解质渗漏和氧化应激。本研究中,通过测定REC和MDA,H2O2和O2-来研究膜渗漏,以阐明由于氧化应激引起的植物损伤。实验结果表明,随时间延长,Cd诱导这些分子的过度积累导致紫花苜蓿种子中的氧化损伤并抑制幼苗生长。生理变化等多种机制都与植物阻止或减轻非生物或非生物胁迫有关。应对环境胁迫,许多植物物中的Pro会积累,Ahmad曾报道了NO在非生物胁迫下Pro的代谢中的调节作用。在本研究中,暴露于Cd胁迫的幼苗Pro水平高于对照幼苗,应用SNP后,其水平进一步下降,特别是当SNP浓度高于200μM时,但长期暴露于200μM以上浓度的SNP时呈现Pro含量增加的趋势,这进一步证实了已有报道的类似的研究结果。为了应对和中和ROS的毒性,植物发展的内源性抗氧化酶系统包括CAT,POD,SOD,APX,GR等抗氧化酶。SOD通过促进超氧化物自由基转化来保护细胞免受氧化应激的最有效的酶被极大地抑制,其活性显著低于对照(CK),可能是由于Zn2+-SOD复合物的氢键弱,固体含量低,Cd2+可以很容易地替代SOD活性位点的Zn2+,使SOD活性降低。在本研究中,Cd的积累导致H2O2和O2-的显著产生,SOD活性降低,而采用外源性NO处理则使SOD活性水平提高,特别是在处理的前3天,处理后期,随SNP浓度的增加,SOD酶活性虽然有所降低但仍然高于或与Cd单独处理持平,证实了早期关于水稻和番茄幼苗研究中类似的报道。外源SNP处理使POD活性水平在整个萌发过程中均高于Cd单独处理,表明POD在萌发过程中都保持较高活性以清除H2O2。APX仅在萌发前2天当SNP浓度低于200μM时活性增强,SNP浓度大于200μM时活性减弱,在萌发后期(3-5天)SNP处理使APX活性显著低于Cd单独处理,且随SNP浓度的增加无显著性变化,表明APX仅在紫花苜蓿种子萌发初期具有清除ROS的作用。CAT与APX类似,在萌发前4天具有清除ROS的作用。Cd胁迫使GR活性从萌发第2天起就显著低于CK,而SNP处理仅在萌发第1天和第3天使GR活性有所升高,但随SNP浓度的增加并没有显示出规律性的变化,表明NO和GR活性诱导之间的关系较为复杂,有待进一步研究。3、外源NO能够增加Cd胁迫下紫花苜蓿幼苗贮藏储备物质的含量,增强相关酶活性。发芽过程的特征是通过储存物质,如碳水化合物和蛋白质等通过水解酶水解新形成的物质用于形成新的结构。储存物质被水解酶水解为呼吸作用和各种合成代谢反应提供能量,以形成代谢产物。负责这些储备物质如淀粉分解的酶是α-淀粉酶和β-淀粉酶,已有报道,Cd胁迫会导致储备物质的动员受到限制。随萌发时间的延长和SNP浓度的增加,SNP激活α-淀粉酶和β-淀粉酶,调节Cd胁迫条件下紫花苜蓿种子中可溶性蛋白、还原糖和可溶性糖含量。30μMCd胁迫使紫花苜蓿种子α和β-淀粉酶活性显著降低,可溶性蛋白和淀粉含量也在处理的第2天显著下降,这是抑制种子萌发的主要因子。本研究表明,外源SNP对30μMCd胁迫下紫花苜蓿幼苗的萌发有很强的刺激作用,使淀粉酶、可溶性蛋白和淀粉含量显著高于仅用Cd处理的幼苗,这一研究结果与外源性NO对Cd胁迫下发芽的豌豆种子淀粉酶活性影响的研究结果一致。通过在Cd处理的幼苗中施用外源NO,α-淀粉酶和β-淀粉酶的活性均显著增强;在处理第2天,可溶性淀粉和蛋白质的含量均高于仅用Cd处理的幼苗中的含量,这有利于改善紫花苜蓿幼苗的萌发。实验的前三天,SNP浓度低于100μM时,能调节可溶性糖的含量,当浓度超过100μM有相反的影响。除了SNP浓度在25和100μM时幼苗还原糖含量比较高,其他处理下幼苗体内还原糖含量均无显著差异,并且还原糖的产生量几乎是恒定的。4、外源NO显著降低紫花苜蓿种子中Cd的积累并增加种子鲜重和干重。Cd作为非必需和有害元素之一,可以通过与Ca2+和K+等植物中的其他微量和大量营养物质竞争来干扰植物体内平衡,甚至也可以通过水来运输。发生这种情况与鲜重和干重物质的减少有关,已有研究报道,水含量受植物中Cd浓度的高度影响。本研究中,只要种子暴露于Cd胁迫下,与CK种子相比,种子的干重和鲜重也随着暴露时间的延长而受到显著的影响。在向日葵和高粱种子中也有类似的研究结果。据报道外源性NO阻碍重金属进入植物细胞壁,或调节植物细胞壁内多余Cd的细胞分布和积累。本研究实验结果与水稻幼苗研究中,水稻叶中汞的吸收和水稻幼苗对铜的摄取的研究结果类似。这表明SNP可能是保护种子免受重金属毒性的合适的化合物之一。30μM的Cd没有显著降低萌发率,但萌发指数,SVI,根长和茎长受Cd毒性影响很大;发现30μM CdCl_2诱导Pro水平升高,REC,MDA以及ROS过度产生,Cd通过诱导抗氧化酶活性降低的能力对紫花苜蓿种子的防御机制有重大的阻碍作用;α-淀粉酶,β-淀粉酶的活性受Cd毒性的显著影响,导致储存物质可溶性蛋白质、可溶性淀粉、可溶性糖和还原糖的损害;Cd也显著影响种子鲜重和干重。外源NO显示能逆转Cd对萌发率、下胚轴长度、根长、发芽指数和幼苗活力指数的负面影响,发现200μM SNP对30μM Cd毒性的缓解作用最有效。外源NO在抵抗ROS过度生成引起的氧化损伤方面是有效的,因为它可以降低发芽紫花苜蓿幼苗的MDA、REC和ROS水平。外源NO也被揭示为能够增强抗氧化系统。外源NO通过调节α和β-淀粉酶的活化调节可溶性淀粉、还原糖和可溶性糖的动员,在抵消Cd毒性引起的贮藏物质的损害方面起主要作用。外源性NO对植物的Cd吸收产生抑制作用,增加干重和鲜重。一般来说,加入200μM的SNP显示为减轻30μM Cd毒性最显著。紫花苜蓿在修复重金属如Cd、Cu、Pb等污染土壤方面的能力非常突出,而在中国特别是西北地区有大面积种植紫花苜蓿作为畜牧业发展的基础,由此可见其在应对逆境环境下的发展前景很广阔。光合作用是植物新陈代谢以生长的基本生理活动,对于NO在Cd胁迫下对紫花苜蓿净光合速率、蒸腾速率、气孔导度、胞间CO2浓度、叶绿素荧光参数以及叶黄素循环是怎样影响的还有待探讨,这是下一步研究的方向和目标。建议进一步采用遗传和蛋白质组学分析及其他生理学方法,以了解外源NO在植物代谢功能中的细节,这将有助于进一步了解这种多面体化合物,用于植物缓解不同种类生物和非生物胁迫,使紫花苜蓿能够更好的应用于重金属污染土壤的植物修复中。
[Abstract]:Cadmium (Cd) is a toxic and non-essential heavy metal for animal and plant tissues. It is very important to explore and study the mechanism of plant resistance to heavy metals and heavy metals in response to heavy metal injuries. Only thoroughly understand the mechanism of plant resistance to heavy metals can we possibly put forward. Effective measures to prevent plants from being harmed by heavy metals and important ways to improve their resistance to heavy metals are beneficial to the control of heavy metal pollution in the soil, the protection of the earth's ecological environment and the production of green and healthy food. Exogenous nitric oxide (NO) is a signaling molecule that regulates abiotic and biological stresses. Species play an important role in physiological responses. How these physiological responses are caused by heavy metal stress signals are transmitted in plants, what role NO plays in them, how to mobilize defense systems and ultimately affect plant growth and development, physiological metabolism and gene expression, and how to regulate plant tolerance to heavy metals through human regulation Some processes in the body process to improve plant tolerance to heavy metals, increase or decrease the uptake of heavy metals, and apply to phytoremediation and agricultural production in heavy metal contaminated areas have not been reported. It is widely distributed in temperate regions of the world and is most suitable for seedling growth. Its extensive roots make it drought-resistant, enhance the activity of degrading bacteria, and promote the biodegradability of several organic pollutants such as polychlorinated biphenyls (PCBs), polycyclic aromatic hydrocarbons (PAH) and trinitrotoluene (TNT), especially Alfalfa is a promising plant for phytoremediation of toxic metals because of its rich vitamin A, C, E, K, minerals and trace elements, rapid growth, well-developed roots, long life span and good resistance. Alfalfa is considered as one of the environmental protection technologies for phytoremediation of heavy metal contaminated soils. In order to study the toxic effects of sodium nitroprusside (SNP, exogenous NO donor) on cadmium during the germination of Alfalfa seedlings, alfalfa has the greatest absorption and enrichment capacity. The effects of exogenous NO on the germination and growth of Alfalfa Seeds under Cd stress were analyzed. The experiment was carried out for 5 days, every 24 hours and recorded and measured. The results showed that the toxicity of CdCl_2 to alfalfa seed germination and seedling growth increased with the increase of CdCl_2 concentration. The growth of alfalfa seedlings was obviously inhibited but not significantly affected by CdCl_2. High concentration of CdCl_2 could lead to a serious decrease in germination rate, thus reducing the available samples of seedlings produced in each group of experiments. Alfalfa seeds were treated with 100,200,300,400 and 500 mu M SNP for 5 days, and the following indices were recorded and measured every 24 hours: germination rate, germination index, seedling vigor index, root length and bud length as germination parameters; proline (Pro) level, relative conductivity (REC), lipid peroxidation (MDA) and reactive oxygen species (ROS), such as H2O2 and O2 - production rate as Cd production. Oxidative stress reaction parameters; superoxide dismutase (SOD), peroxidase (POD), ascorbic acid peroxidase (APX), catalase (CAT), glutathione reductase (GR) and other antioxidant enzymes as antioxidant mechanism parameters; alpha-amylase and beta-amylase as hydrolytic enzyme parameters; soluble protein, soluble protein, soluble amylase; Starch, soluble sugar and reducing sugar were used as storage material parameters. 1. Exogenous NO could alleviate the inhibition of Cd stress on Seed Germination and seedling growth of alfalfa. SNP showed a significant role in alleviating heavy metal stress and promoting plant growth. At the optimum concentration, the alleviation increased linearly, while above the optimum concentration, plant growth was inhibited. This response was more likely to be the result of effective exogenous NO activating plant defense or tolerance mechanisms. Fruit. This confirms previous studies on the protection of nitric oxide during seed germination under arsenic stress. Studies on Rice Seed Germination and seedling growth under cadmium stress have clarified the importance of exogenous nitric oxide to cadmium toxicity. SNP can significantly improve germination rate, germination index and seedling vigor index, and has been reported in sesame seedlings. In this study, exogenous nitric oxide successfully alleviated Cd damage and probably operated in a similar manner to other biotic or abiotic stresses. However, above the optimal concentration of SNP, plants or seedlings exhibited the same defense mechanism similar to that of perennial ryegrass under Cd stress. 200 mu SNP could effectively alleviate Cd stress. Understanding the inhibitory effect of 30 mu cadmium on Alfalfa Seed Germination and growth. 2. Exogenous NO can alleviate the oxidative damage of Alfalfa under Cd stress by activating the antioxidant system, scavenging reactive oxygen species. Cd can induce ROS in seedlings, such as O2 - and H2O2, which lead to membrane lipid peroxidation, electrolyte leakage and oxidative stress. EC and MDA, H2O2 and O2 - were used to study membrane leakage in order to elucidate the plant damage caused by oxidative stress. The results showed that over time, excessive accumulation of these molecules induced by Cd led to oxidative damage in alfalfa seeds and inhibited seedling growth. Physiological changes and other mechanisms involved in plant inhibition or mitigation of abiotic or abiotic stress. In this study, the level of Pro in seedlings exposed to Cd stress was higher than that in the control seedlings. The level of Pro in seedlings exposed to Cd stress decreased further after the application of SNP, especially when the concentration of SNP was higher than 200 mu M, but long-term exposure to 20 mu. In response to and neutralizing ROS toxicity, endogenous antioxidant enzymes developed in plants include CAT, POD, SOD, APX, GR and other antioxidant enzymes. SOD protects cells from oxidation by promoting the transformation of superoxide free radicals. The most effective enzyme under stress was significantly inhibited, and its activity was significantly lower than that of control (CK), possibly because of the weak hydrogen bond of Zn2 + - SOD complex and the low solid content. Cd2 + could easily replace Zn2 + of SOD active site, so that the activity of SOD decreased. In this study, the accumulation of Cd LED to the significant production of H2O2 and O2 - and the decrease of SOD activity. The activity of SOD was increased by NO treatment, especially in the first three days after treatment. With the increase of SNP concentration, the activity of SOD was still higher than or equal to that of Cd treatment alone, which confirmed similar reports in early studies on rice and tomato seedlings. The activity of POD was higher than that of Cd alone, indicating that POD maintained higher activity in germination to scavenge H2O2. APX increased only 2 days before germination when SNP concentration was lower than 200 mu M, and decreased when SNP concentration was higher than 200 mu M. SNP treatment made APX activity significantly lower than that of Cd alone in late germination (3-5 days), and there was no significant change with the increase of SNP concentration. The results showed that APX scavenged ROS only at the early germination stage of Alfalfa seeds. CAT and APX scavenged ROS 4 days before germination. Cd stress made GR activity significantly lower than CK from the 2nd day of germination, while SNP treatment only increased GR activity on the 1st and 3rd day of germination, but showed no regularity with the increase of SNP concentration. 3. Exogenous NO can increase the content of storage substances and enhance the activity of related enzymes in Alfalfa Seedlings under Cd stress. The germination process is characterized by the hydrolysis of storage substances, such as carbohydrates and proteins, by hydrolases. Storage substances are hydrolyzed by hydrolases to provide energy for respiration and various anabolic reactions to form metabolites. The enzymes responsible for these storage substances, such as starch decomposition, are alpha-amylase and beta-amylase. It has been reported that Cd stress can limit the mobilization of reserve substances. The soluble protein, reducing sugar and soluble sugar content of alfalfa seeds were regulated by SNP activating alpha-amylase and beta-amylase. The activity of alpha-amylase and beta-amylase in alfalfa seeds were significantly decreased by 30 mu MCd stress, and the content of soluble protein and starch were also significantly decreased by the second day of treatment. The results showed that exogenous SNP could stimulate the germination of Alfalfa Seedlings under 30 mu MCd stress, and the contents of amylase, soluble protein and starch were significantly higher than those of alfalfa seedlings treated only with Cd. The results showed that exogenous NO could affect the amylase activity of pea seeds under Cd stress. The activity of exogenous NO, alpha-amylase and beta-amylase in seedlings treated with Cd increased significantly, and the contents of soluble starch and protein in seedlings treated with Cd were higher than those in seedlings treated only with Cd on the second day, which was beneficial to improving the germination of Alfalfa seedlings. The content of reducing sugar in seedlings was not significantly different under other treatments except SNP concentration at 25 and 100 mu M, and the production of reducing sugar was almost constant. 4. Exogenous NO significantly reduced the accumulation of Cd in alfalfa seeds and decreased the accumulation of Cd in alfalfa seeds. Cd, as one of the non-essential and harmful elements, can interfere with the balance of plants by competing with other trace and large amounts of nutrients in plants such as Ca2+ and K +, and can even be transported by water. This is related to the reduction of fresh and dry weight. Studies have reported that water content is being planted. In this study, as long as the seeds were exposed to Cd stress, the dry weight and fresh weight of the seeds were significantly affected as compared with CK seeds. Similar results were found in sunflower and sorghum seeds. Exogenous NO was reported to block the entry of heavy metals into plant cell walls, or to regulate them. Distribution and accumulation of excess Cd in plant cell wall. The results of this study were similar to those of rice seedling studies. It suggested that SNP might be one of the suitable compounds to protect seeds from heavy metal toxicity. 30 mu Cd did not significantly reduce the germination rate, but germinated. Hair index, SVI, root length and stem length were significantly affected by Cd toxicity; it was found that 30 mu CdCl_2 could induce the increase of Pro level, the excessive production of REC, MDA and ROS, and Cd could significantly inhibit the defense mechanism of alfalfa seeds by inducing the decrease of antioxidant enzyme activity; the activities of alpha-amylase and beta-amylase were significantly affected by Cd toxicity, resulting in the overproduction of REC, MDA and ROS. Soluble proteins, starches, sugars and reducing sugars were damaged by storage substances, and Cd also significantly affected the fresh and dry weight of seeds.
【学位授予单位】：兰州交通大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：X173
本文编号：2250280
[Abstract]:Cadmium (Cd) is a toxic and non-essential heavy metal for animal and plant tissues. It is very important to explore and study the mechanism of plant resistance to heavy metals and heavy metals in response to heavy metal injuries. Only thoroughly understand the mechanism of plant resistance to heavy metals can we possibly put forward. Effective measures to prevent plants from being harmed by heavy metals and important ways to improve their resistance to heavy metals are beneficial to the control of heavy metal pollution in the soil, the protection of the earth's ecological environment and the production of green and healthy food. Exogenous nitric oxide (NO) is a signaling molecule that regulates abiotic and biological stresses. Species play an important role in physiological responses. How these physiological responses are caused by heavy metal stress signals are transmitted in plants, what role NO plays in them, how to mobilize defense systems and ultimately affect plant growth and development, physiological metabolism and gene expression, and how to regulate plant tolerance to heavy metals through human regulation Some processes in the body process to improve plant tolerance to heavy metals, increase or decrease the uptake of heavy metals, and apply to phytoremediation and agricultural production in heavy metal contaminated areas have not been reported. It is widely distributed in temperate regions of the world and is most suitable for seedling growth. Its extensive roots make it drought-resistant, enhance the activity of degrading bacteria, and promote the biodegradability of several organic pollutants such as polychlorinated biphenyls (PCBs), polycyclic aromatic hydrocarbons (PAH) and trinitrotoluene (TNT), especially Alfalfa is a promising plant for phytoremediation of toxic metals because of its rich vitamin A, C, E, K, minerals and trace elements, rapid growth, well-developed roots, long life span and good resistance. Alfalfa is considered as one of the environmental protection technologies for phytoremediation of heavy metal contaminated soils. In order to study the toxic effects of sodium nitroprusside (SNP, exogenous NO donor) on cadmium during the germination of Alfalfa seedlings, alfalfa has the greatest absorption and enrichment capacity. The effects of exogenous NO on the germination and growth of Alfalfa Seeds under Cd stress were analyzed. The experiment was carried out for 5 days, every 24 hours and recorded and measured. The results showed that the toxicity of CdCl_2 to alfalfa seed germination and seedling growth increased with the increase of CdCl_2 concentration. The growth of alfalfa seedlings was obviously inhibited but not significantly affected by CdCl_2. High concentration of CdCl_2 could lead to a serious decrease in germination rate, thus reducing the available samples of seedlings produced in each group of experiments. Alfalfa seeds were treated with 100,200,300,400 and 500 mu M SNP for 5 days, and the following indices were recorded and measured every 24 hours: germination rate, germination index, seedling vigor index, root length and bud length as germination parameters; proline (Pro) level, relative conductivity (REC), lipid peroxidation (MDA) and reactive oxygen species (ROS), such as H2O2 and O2 - production rate as Cd production. Oxidative stress reaction parameters; superoxide dismutase (SOD), peroxidase (POD), ascorbic acid peroxidase (APX), catalase (CAT), glutathione reductase (GR) and other antioxidant enzymes as antioxidant mechanism parameters; alpha-amylase and beta-amylase as hydrolytic enzyme parameters; soluble protein, soluble protein, soluble amylase; Starch, soluble sugar and reducing sugar were used as storage material parameters. 1. Exogenous NO could alleviate the inhibition of Cd stress on Seed Germination and seedling growth of alfalfa. SNP showed a significant role in alleviating heavy metal stress and promoting plant growth. At the optimum concentration, the alleviation increased linearly, while above the optimum concentration, plant growth was inhibited. This response was more likely to be the result of effective exogenous NO activating plant defense or tolerance mechanisms. Fruit. This confirms previous studies on the protection of nitric oxide during seed germination under arsenic stress. Studies on Rice Seed Germination and seedling growth under cadmium stress have clarified the importance of exogenous nitric oxide to cadmium toxicity. SNP can significantly improve germination rate, germination index and seedling vigor index, and has been reported in sesame seedlings. In this study, exogenous nitric oxide successfully alleviated Cd damage and probably operated in a similar manner to other biotic or abiotic stresses. However, above the optimal concentration of SNP, plants or seedlings exhibited the same defense mechanism similar to that of perennial ryegrass under Cd stress. 200 mu SNP could effectively alleviate Cd stress. Understanding the inhibitory effect of 30 mu cadmium on Alfalfa Seed Germination and growth. 2. Exogenous NO can alleviate the oxidative damage of Alfalfa under Cd stress by activating the antioxidant system, scavenging reactive oxygen species. Cd can induce ROS in seedlings, such as O2 - and H2O2, which lead to membrane lipid peroxidation, electrolyte leakage and oxidative stress. EC and MDA, H2O2 and O2 - were used to study membrane leakage in order to elucidate the plant damage caused by oxidative stress. The results showed that over time, excessive accumulation of these molecules induced by Cd led to oxidative damage in alfalfa seeds and inhibited seedling growth. Physiological changes and other mechanisms involved in plant inhibition or mitigation of abiotic or abiotic stress. In this study, the level of Pro in seedlings exposed to Cd stress was higher than that in the control seedlings. The level of Pro in seedlings exposed to Cd stress decreased further after the application of SNP, especially when the concentration of SNP was higher than 200 mu M, but long-term exposure to 20 mu. In response to and neutralizing ROS toxicity, endogenous antioxidant enzymes developed in plants include CAT, POD, SOD, APX, GR and other antioxidant enzymes. SOD protects cells from oxidation by promoting the transformation of superoxide free radicals. The most effective enzyme under stress was significantly inhibited, and its activity was significantly lower than that of control (CK), possibly because of the weak hydrogen bond of Zn2 + - SOD complex and the low solid content. Cd2 + could easily replace Zn2 + of SOD active site, so that the activity of SOD decreased. In this study, the accumulation of Cd LED to the significant production of H2O2 and O2 - and the decrease of SOD activity. The activity of SOD was increased by NO treatment, especially in the first three days after treatment. With the increase of SNP concentration, the activity of SOD was still higher than or equal to that of Cd treatment alone, which confirmed similar reports in early studies on rice and tomato seedlings. The activity of POD was higher than that of Cd alone, indicating that POD maintained higher activity in germination to scavenge H2O2. APX increased only 2 days before germination when SNP concentration was lower than 200 mu M, and decreased when SNP concentration was higher than 200 mu M. SNP treatment made APX activity significantly lower than that of Cd alone in late germination (3-5 days), and there was no significant change with the increase of SNP concentration. The results showed that APX scavenged ROS only at the early germination stage of Alfalfa seeds. CAT and APX scavenged ROS 4 days before germination. Cd stress made GR activity significantly lower than CK from the 2nd day of germination, while SNP treatment only increased GR activity on the 1st and 3rd day of germination, but showed no regularity with the increase of SNP concentration. 3. Exogenous NO can increase the content of storage substances and enhance the activity of related enzymes in Alfalfa Seedlings under Cd stress. The germination process is characterized by the hydrolysis of storage substances, such as carbohydrates and proteins, by hydrolases. Storage substances are hydrolyzed by hydrolases to provide energy for respiration and various anabolic reactions to form metabolites. The enzymes responsible for these storage substances, such as starch decomposition, are alpha-amylase and beta-amylase. It has been reported that Cd stress can limit the mobilization of reserve substances. The soluble protein, reducing sugar and soluble sugar content of alfalfa seeds were regulated by SNP activating alpha-amylase and beta-amylase. The activity of alpha-amylase and beta-amylase in alfalfa seeds were significantly decreased by 30 mu MCd stress, and the content of soluble protein and starch were also significantly decreased by the second day of treatment. The results showed that exogenous SNP could stimulate the germination of Alfalfa Seedlings under 30 mu MCd stress, and the contents of amylase, soluble protein and starch were significantly higher than those of alfalfa seedlings treated only with Cd. The results showed that exogenous NO could affect the amylase activity of pea seeds under Cd stress. The activity of exogenous NO, alpha-amylase and beta-amylase in seedlings treated with Cd increased significantly, and the contents of soluble starch and protein in seedlings treated with Cd were higher than those in seedlings treated only with Cd on the second day, which was beneficial to improving the germination of Alfalfa seedlings. The content of reducing sugar in seedlings was not significantly different under other treatments except SNP concentration at 25 and 100 mu M, and the production of reducing sugar was almost constant. 4. Exogenous NO significantly reduced the accumulation of Cd in alfalfa seeds and decreased the accumulation of Cd in alfalfa seeds. Cd, as one of the non-essential and harmful elements, can interfere with the balance of plants by competing with other trace and large amounts of nutrients in plants such as Ca2+ and K +, and can even be transported by water. This is related to the reduction of fresh and dry weight. Studies have reported that water content is being planted. In this study, as long as the seeds were exposed to Cd stress, the dry weight and fresh weight of the seeds were significantly affected as compared with CK seeds. Similar results were found in sunflower and sorghum seeds. Exogenous NO was reported to block the entry of heavy metals into plant cell walls, or to regulate them. Distribution and accumulation of excess Cd in plant cell wall. The results of this study were similar to those of rice seedling studies. It suggested that SNP might be one of the suitable compounds to protect seeds from heavy metal toxicity. 30 mu Cd did not significantly reduce the germination rate, but germinated. Hair index, SVI, root length and stem length were significantly affected by Cd toxicity; it was found that 30 mu CdCl_2 could induce the increase of Pro level, the excessive production of REC, MDA and ROS, and Cd could significantly inhibit the defense mechanism of alfalfa seeds by inducing the decrease of antioxidant enzyme activity; the activities of alpha-amylase and beta-amylase were significantly affected by Cd toxicity, resulting in the overproduction of REC, MDA and ROS. Soluble proteins, starches, sugars and reducing sugars were damaged by storage substances, and Cd also significantly affected the fresh and dry weight of seeds.
【学位授予单位】：兰州交通大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：X173
【参考文献】
相关期刊论文 前3条
1 Mehar Fatma;Asim Masood;Tasir S.Per;Faisal Rasheed;Nafees A.Khan;;Interplay between nitric oxide and sulfur assimilation in salt tolerance in plants[J];The Crop Journal;2016年03期
2 ;Effects of cadmium stress on seed germination and seedling growth of Elymus dahuricus infected with the Neotyphodium endophyte[J];Science China(Life Sciences);2012年09期
3 ;Effects of Cadmium Stress on Seed Germination, Seedling Growth and Seed Amylase Activities in Rice (Oryza sativa)[J];Rice Science;2008年04期
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