【作者】 庞彩红；

【导师】 王宝山；

【作者基本信息】 山东师范大学 ， 植物学， 2004， 硕士

【摘要】 越来越多的实验证明，盐胁迫诱导植物体内活性氧（O₂、H₂O₂、OH^-等）产生的增加，引起氧化胁迫。其中O₂^-和H₂O₂在盐对植物的伤害机制中起着主要作用。O₂^-主要由超氧化物歧化酶（SOD）通过歧化反应形成H₂O₂和O₂而进行清除，SOD在降低O₂^-毒害的同时却增加了细胞内H₂O₂的含量，除此之外细胞内一些酶促或非酶促反应也能产生H₂O₂，H₂O₂在过渡金属存在下可转化为氧化活性更高的OH·。因此，植物能否有效地清除逆境下产生的H₂O₂对其抗逆性至关重要。本实验比较系统地研究了不同浓度NaCl处理下，盐地碱蓬叶片中清除H₂O₂的各种酶（CAT、GR、GST）的活性及非酶类物质（GSH）含量的变化情况及H₂O₂水平，并结合其它生理指标阐明他们在盐地碱蓬耐盐性中的作用，旨在了解盐地碱蓬的耐盐性与H₂O₂清除系统的关系，进一步完善盐生植物的耐盐机理，结果如下： 1、NaCl处理对盐地碱蓬生长状况的影响 盐地碱蓬整株鲜重随NaCl处理浓度的增加出现先增后降的趋势，在200mmol／L，整株鲜重与对照的差异达到极显著水平（P＜0.01），当盐浓度达到400mmol／L时鲜重有所下降，但仍显著高于对照，这说明一定的盐浓度NaGl会促进盐地碱蓬的生长，当环境中盐分浓度达到一定浓度后才会对植物生长产生抑制。 盐地碱蓬叶片中Na⁺、Cl^-离子含量随着盐处理浓度的升高而增大，而且表现出基本相同的增加趋势，而K⁺离子的含量随着盐处理浓度的升高而下降。盐地碱蓬叶片细胞汁液渗透势的变化与NaCl处理条件下Na⁺、Cl^-离子含量的变化相一致，这说明Na⁺、Cl^-在调节盐地碱蓬细胞渗透势过程中起重要作用。渗透势都随盐处理浓度的增高而降低，有利于细胞从外界吸水，从而保证了体内水分的充分供应。 2、NaCl处理对盐地碱蓬光合特性的效应 盐地碱蓬在NaCl胁迫下，Gs和Ci无明显变化，Pn有增高的趋势，在200mmol／L NaCl条件下Pn达到最大值，这与盐地碱蓬的生长趋势相一致；Pn的升高可能是chla／chlb和ΦPS Ⅱ的增加引起的。 3、NaCl处理对盐地碱蓬细胞膜的伤害 活性氧导致膜脂过氧化，产生膜伤害。由于膜脂过氧化是最主要的氧化伤害，所以用MDA来表示氧化伤害的程度。如图16所示，盐胁迫下，盐地碱蓬MDA含量增加幅度小（P＞0.05），膜透性改变不明显（图17），说明盐地碱蓬膜必承蹄i曹人.学硕去穆犷羌{竺‘论丈2004伞甲刀系统受损伤较轻，甚至不会对植物生长产生影响，意味着盐地碱蓬具有很强的抵抗盐胁迫诱导的氧化胁迫能力。 4、NaCI处理对盐地碱蓬叶片中H20:含量的影响 盐地碱蓬叶片中HZO:的含量出现先降后增趋势，但均低于对照，HZOZ含量降低，减小了其对细胞的氧化伤害，200 mmol几处理时差异达到了极显著水平，在此浓度一「盐地碱蓬的生长状况也是最好的。 5、NaCI处理对盐地碱蓬叶片CA飞，的效应 随着欲处理浓度的增加，叶片中CAT的活性呈上升趋势（图19A）。盐地碱蓬叶片C兀1、活性在小于200^ol儿的情况下，随盐处理浓度的增加CAT活性增加并不显著（P>0.05），当浓度增加到200^ol/L时，活性增加迅速（P<0.05），400^ol/L时，活性显著高于对照（P<0.01）。NaCI处理仅诱导CAf酶活性的升高，并没有诱导新的同工酶产生。 6、NaCI处理对盐地碱蓬GR、GSH的效应 不同浓度NaCI处理下，盐地碱蓬叶片中GR、GSH的变化趋势基本相同，均呈先上升后下降的趋势，但与对照相比都达到了显著差异，200^ol/L时GR的活性及GSH的含量最高，与对照相比差异达到了极显著的水平（P<0.01）。 叶绿体中GR主要分布于类囊体膜和叶绿体基质中，其中叶绿体基质中的GR占主导地位。随盐处理浓度的增加，叶绿体中总GR、类囊体膜GR、基质GR的活性呈先上升后下降的趋势，但与对照相比差异显著，200 nn刀nol/L时差异达到了极显著的水平（P<0.01）。这种变化趋势与整个细胞中GR活性的变化趋势是一致的。叶绿体中GR约占整个细胞中可溶性GR的75%左右，由此可推测叶绿体中的ASA一GSH循环在整个细胞中占主导地位，在清除叶绿体中的HZOZ时起主要作用。 非变性电泳显示盐地碱蓬叶片中GR同工酶只有一种，活性受到NaCI处理的促进，但在400 mmol几NaCI时有所下降。 7、NaCI处理对盐地碱蓬叶片GST的效应 不同浓度NaCI处理下，盐地碱蓬叶片中GST也具有先上升后下降的趋势，有所不同的是GST只在200 mm0UL时与对照相比达到了显著差异的水平 (P（0 .05）。 用Northem印迹杂交法分析盐地碱蓬GST基因的表达特性，结果表明，400mmol几的NaCI处理对盐地碱蓬GS丁基因的诱导先是受抑制，然后显著增强，NaCI溶液处理48h的mRNA水平约为对照植株的2一3倍。 总之，盐地碱蓬具有很强的适应盐渍环境的能力和抗氧化胁迫能力。在高盐胁迫下，盐地碱蓬仍能生长良好，光合作用依然高效地进行，HZO:水平不但功乐…顺应久.贷硬去返不无…全讨丈2004卒4刀没上升，反而下降，M以含量也无显著变化，这与其具有高效的叶绿体HZO:清除系统相一致，这可能是盐生植物l耐盐的重要机制之一。更多还原

【Abstract】 The enhanced production of ROS during salt stress can pose oxidative stress to cells and lead to the oxidative destruction. High NaCl concentrations seem to impair electron transport and lead to the formation of ROS. Plants have well-developed antioxidant defense systems against ROS, involving both enzymatic and non-enzymatic mechanisms. The chief toxicity of O2 and H2O2 is thought to reside in their ability to initiate cascade reactions that result in the production of the hydroxyl radical and other destruction species such as lipid peroxides. These dangerous cascades are prevented by efficient operation of the cell’s antioxidant defenses. SOD converts superoxide radicals (02) to hydrogen peroxide (H2O2). Hydrogen peroxide acts as a precursor of more cytotoxic or highly reactive oxygen derivatives, such as peroxynitrite or HO .It is very important to scavenge the excess H2O2. CAT GR and GST are related to H2O2-scavenging. In the present study, salt-dilution halophyte S. salsa grown under different salt concentrations was used to investigate the activities of H2O2-scavenging enzymes. The main results are as follows.1. Effects of NaCI treatment on growth of S. salsa seedlings.The growth of S. salsa was significantly enhanced by NaCl treatment. The fresh weight increased significantly with the increase of NaCl concentration up to 200 mmol/L and decreased slightly at higher concentrations. However, the fresh weight of the seedlings treated with 400 mmol/L NaCl was still higher than that of controls. Under the condition of 200 mmol/L NaCl, the fresh weight of 5. salsa reached the maximum.S. salsa plants accumulated high Na+ in salt environment. The leaves of S. salsa are the main organs of Na+accumulation. Leaf Na+ and Cl" contents were significantly increased with the increase of NaCl concentration (0-400 mmol/L), whereas K+ content and osmotic potential of the cell sap of the leaves were drastically decreased. Since the growth of S. salsa seedlings was stimulated by salt stress, the decrease of K+ concentration in the cell sap in a certain extent does not affect the growth of S. salsa. S. salsa modulate the osmolarity by absorbing abundant Na+. 2. Effects of salt-treatment on the photosynthetic characters of S. salsa seedlings.Gs and Ci of S. salsa seedlings did not show responses to salt treatment. Pn andPS II increased with the increase of NaC! concentration. Under 200 mmol/L NaCI, Pn and0 PS II of 5. salsa seedlings reached the maximum.In S. salsa seedlings, chla+b was not affected by salt stress (0-400 mmol/L), chla/b was significantly increased under conditions of 200 mmol/L NaCI (up to 171.1%), then decreased by 400 mmol/L NaCI (up to 136.2%).3. Radical damage to 5. salsa seedlings.lipid peroxidation (MDA) was used as indicators of NaCI stress to test the radical damage. In low NaCI concentration (0-200 mmol/L), leaf MDA did not show distinct increase. Under the condition of 400 mmol/L NaCI, leaf MDA was only increased by1 7.8%.4. Leaf CAT and isoforms of S. salsa response to salt treatment.In S. salsa leaves three isoforms of CAT were detected after staining of the native gels. Salt stress did not affect isoform pattern of CAT. The activity of CAT increased with the increase of NaCI concentration. In low NaCI concentration (0-200 mmol/L), activity of CAT did not show distinct increase, while under the condition of 200-400 mmol/L NaCI, CAT activity increased significantly.5. Effects of NaCltreatment on activity of GR GSH and GR isoforms of S.salsaSalt stress did not change isoform pattern of GR, just increased its activity. There was only one GR isoforms in S. salsa seedlings.GR and GSH belong to ascorbate-glutathione cycle. Under NaCI treatment, the activity of GR increased with the increase of NaCI concentration up to 200 mmol/L and decreased slightly at higher concentration. However, GR activity of NaCl-treated seedlings was still significantly higher than that of control plants .The content of GSH exhibited same trend to GR’sUnder salt stress, similar changes were observ更多还原