超表达拟南芥2-烯醛还原酶基因对烟草抗旱性的作用机理分析

为研究是否可以利用2-烯醛还原酶(AER)来清除活性氧下游的醛自由基达到提高植物的抗旱性,以超表达拟南芥AER基因烟草和野生型烟草(SR)为研究材料,利用干旱胁迫处理进行抗旱性分析,测定了干旱胁迫及复水后各个烟草株系的生物量、光合速率、叶绿素荧光参数、叶绿素含量、MDA和H2O2含量等指标。结果显示:(1)干旱胁迫下,转基因烟草株系的生物量、叶绿素含量、净光合速率、PSⅡ最大光化学效率及H2O2的清除能力均显著高于对照;(2)复水之后,烟草植株的各项生理指标都得到一定程度的恢复,而转基因株系相比于野生型恢复迅速,恢复能力更强。研究认为,超表达AER基因可以通过清除活性氧及其下游醛自由基来提高烟草的抗旱能力。     

甲基紫精胁迫下转TheIF1A基因烟草的活性氧代谢

真核翻译起始因子e IF1家族基因具有一定的抗逆调节能力。前期研究表明柽柳The IF1A基因能提高转基因植物的抗旱耐盐胁迫能力。本研究旨在对该基因的抗氧化能力进行分析,探讨The IF1A基因是否具有抗氧化能力。组织化学染色结果显示,甲基紫精胁迫下转The IF1A基因烟草叶片、保卫细胞、根尖积累的活性氧明显少于野生型烟草(WT),H2O2含量测定结果也表明转基因株系的H_2O_2含量显著低于非转基因株系。此外,甲基紫精胁迫下转The IF1A基因烟草的CAT活性、POD活性显著高于WT株系,表明过表达The IF1A可能通过提高保护酶活性来调节体内活性氧清除能力进而改善植株活性氧积累,提高抗氧化能力。     

红光与远红光比值对盐胁迫下番茄幼苗抗氧化能力的影响

该试验以‘Money Maker’野生型(CK)和以‘Money Maker’为背景的光敏色素B1突变体(phyB1)番茄植株为试材,分析红光与远红光比值(R∶FR)分别为7.4、1.2和0.8的光环境下,受盐胁迫(100mmol·L-1 NaCl)的番茄幼苗根系形态指标及叶和根中渗透调节物质含量、活性氧含量、抗氧化酶活性、MDA含量和相对电解质渗透率的变化,以解释不同R∶FR值对番茄幼苗抗氧化能力的影响以及光敏色素B1在其中的作用。结果显示:(1)在盐胁迫条件下,野生型和突变体phyB1番茄幼苗的根系形态指标(总根长、总根表面积、根尖数、根叉数)比对照显著降低,叶和根中渗透调节物质(可溶性蛋白和脯氨酸)含量、活性氧(O-·2和H2O2)含量、抗氧化酶(SOD、POD、CAT)活性、MDA含量和相对电解质渗透率均比对照显著升高。(2)在盐胁迫条件下降低植株生长光环境中的R∶FR值,野生型番茄植株的根系形态指标以及叶和根中渗透调节物质含量及抗氧化酶活性均比盐胁迫下显著升高,叶和根中活性氧含量、MDA含量和相对电解质渗透率均比盐胁迫下显著降低,而在phyB1突变体番茄植株中,以上指标在不同R∶FR值处理间均没有显著变化。研究发现,低R∶FR值能够促进野生型番茄根系的生长,提高叶和根中渗透调节物质含量和抗氧化酶活性,降低叶和根中活性氧含量、MDA含量和相对电解质渗透率,增强野生型番茄幼苗的抗氧化能力,促进番茄幼苗的生长,且当R∶FR值为0.8时整体效果最优;光敏色素B1在低R∶FR值促进番茄幼苗抗氧化能力中发挥了重要作用。     

番茄转录辅激活子基因LeMBF1在非生物胁迫中的抗性分析

为了研究番茄转录辅激活子基因LeMBF1在非生物胁迫中的作用,以LeMBF1超表达转基因番茄和野生型番茄组培苗为材料,对其进行高温、机械损伤、低温、H2O2、干旱和ABA等非生物胁迫处理.半定量RT-PCR分析表明,LeMBF1在根、幼叶、成熟叶片、花和果实的不同时期均有表达,但在幼叶中表达量最高,成熟叶片和果实中表达量较低.高温处理后,下游防卫反应基因HSP90、Apx2、Zat、PR1在超表达番茄植株中的表达量明显高于野生型番茄植株,并且LeMBF1超表达番茄植株存活率较高.机械损伤可以明显的诱导LeMBF1基因表达;H2O2、干旱、低温、和ABA处理后,对下游防卫基因的表达量有不同程度的影响.以上结果表明LeMBF1基因参与了多种植物非生物胁迫防御反应,并对提高番茄的抗性有一定的作用.     

过表达LeNLP4转录因子提高番茄抗低温胁迫能力

NAC（NAM-ATAF1,2-CUC2）转录因子在植物胁迫响应中起重要作用。为了探讨三舭丹基因在番茄抗低温胁迫中的功能,分离了番茄LeNLP4转录因子基因,并获得转正义LeNLP4基因番茄植株。荧光定量PCR分析表明,LeNLP4的表达受低温诱导。与野生型植株相比,在4℃胁迫下转基因植株具有较高的生长量和光系统II（PSH）最大光化学效率（Fv／Fm）、过氧化氢（H2O2）和超氧阴离子（O2-）清除速率、抗坏血酸过氧化物酶（APX）和超氧化物歧化酶（SOD）活性,以及较低的丙二醛（MDA）含量和相对电导率（REC）。过表达株系中SICBF1的表达高于野生型。上述结果表明,LeNLP4的过表达提高了转基因番茄抗低温胁迫能力。     

陆地棉GhCDPK1基因响应干旱胁迫的功能初探

钙依赖性蛋白激酶(CDPKs)是一类重要的钙信号感受蛋白和响应蛋白,在植物干旱、低温、盐碱等非生物胁迫应答中起着重要的调控作用。为探讨陆地棉GhCDPK1基因在干旱胁迫下所起的作用,该研究利用实时荧光定量PCR技术分析了PEG模拟干旱胁迫下该基因的表达量,发现GhCDPK1基因受干旱胁迫诱导。通过构建植物表达载体pCAMBIA2300-GhCDPK1,采用农杆菌介导的叶盘法转化模式植物烟草,发现干旱胁迫下转基因植株保水能力明显高于野生型植株,叶绿素、脯氨酸、可溶性蛋白含量及POD、SOD活性也高于野生型植株,而丙二醛含量低于野生型植株。研究结果表明,GhCDPK1基因作为正向调控因子响应干旱胁迫诱导,过表达GhCDPK1基因可以使植株积累更多的渗透调节物质、增强抗氧化系统酶的活性和维持细胞膜的稳定性来提高植物抵御外界干旱胁迫的能力。     

6种草本植物对干旱胁迫和CO2浓度升高交互作用的生长响应

以CO2浓度升高为主要标志的全球气候变化及由其引起的极端气候变化对陆地生态系统产生了重要的影响。利用步入式CO2生长室模拟研究了CO2浓度变化(400和700μL/L)和干旱胁迫(水分充足CK:100%FC(田间持水量);中度干旱MS:40%FC;重度干旱SS:20%FC)的交互作用对草本植物网果酸模(Rumex chalepensis)、野豌豆(Vicia sepium)、泥胡菜(Hemmistepta lyrata)、风轮菜(Clinopodium chinense)、藜(Chenopodium album)和玉米石(Sedum album)生长特性的影响。结果表明:CO2浓度升高总体上刺激了网果酸模、野豌豆、泥胡菜、风轮菜和藜这5种C3植物在任何水分条件下的生长,也刺激了玉米石在水分条件较好下的生长;干旱胁迫总体上抑制了所有6种植物的生长,但中度干旱胁迫有刺激CAM植物玉米石生长的趋势。CO2浓度升高能否缓解干旱的负面影响具有明显的种间差异:CO2浓度升高减缓了干旱胁迫对泥胡菜和风轮菜的负面影响,这种缓解作用在网果酸模和野豌豆中显著降低,对藜没有明显的促进作用,对干旱下的玉米石的生长却起到了抑制作用。CO2浓度升高总体上增加了根质量分数和干物质含量;干旱胁迫明显提高了6种草本植物的根生物量的分配比例,降低了干物质含量;但CO2浓度升高和干旱胁迫的交互作用可导致不同的物种产生不同的响应,说明植物能够通过调节生物量分配和植株本身的水分含量保持能力来适应CO2浓度和干旱胁迫的交互影响,这种调节能力取决于植物在碳的吸收和水分散失之间的平衡"trade-off"。研究结果有助于增进草本植物对未来气候变化的适应性理解,为评估和预测全球气候和水文变化对植物的生理生态影响提供理论依据。     

拟南芥AtCIPK23基因对烟草抗旱能力的影响

为了探索拟南芥AtCIPK23基因对烟草耐旱能力的影响,对3个转AtCIPK23基因阳性纯合株系KA13、KA14和KA44与野生型烟草K326（对照）进行了自然干旱处理,测定离体叶片的失水速率、叶绿素含量、相对电导率、脯氨酸和可溶性糖含量,并分析了转基因及野生型材料对活性氧的清除能力,对活性氧清除基因NtSOD、NtCAT和NtAPX及干旱胁迫相关基因NtDREB、NtLEA5和NtCDPK2的表达量进行检测。结果表明：（1）转基因烟草离体叶片的失水速率明显低于K326;自然干旱7 d后,野生型K326出现了明显的干旱胁迫症状;干旱7 d进行复水后,转基因株系的复水存活率明显高于K326。（2）转基因株系中的叶绿素、脯氨酸及可溶性糖含量比K326显著提高,电导率则明显降低。（3）野生型烟草K326中H₂O₂的积累量明显高于3个转基因株系,转基因株系中ROS清除机制的3个关键基因NtSOD、NtCAT和NtAPX被诱导上调表达。（4）抗旱相关基因NtDREB、NtLEA5和NtCDPK2仅在转基因烟草中受干旱诱导。研究认为,AtCIPK23基因可能具有提高植物抗旱能力的功能。     

拟南芥MT-Ⅱ过量表达提高抗旱性

富含巯基的植物Ⅱ型金属硫蛋白(MT)对植物抵抗重金属胁迫具有重要作用,其中一个可能机制是金属硫蛋白可能猝灭重金属引起的氧化胁迫.利用转MT-Ⅱ基因和野生型拟南芥(Arabidopsis thaliana)植株来对比研究MT在胁迫过程中通过清除氧自由基,特别是H2O2而对植物抗旱性的影响.研究表明,转基因型拟南芥能有效维持体内氧化-还原势,减少MDA的产生,从而缓解干旱胁迫引起的伤害,提高抗旱性.     

拟南芥CBF2基因的抗旱性研究

以拟南芥为材料,根据已报道的序列设计引物克隆了CBF2基因及rd29A基因启动子,并构建了植物表达载体pBI-rd-cbf,用以转化烟草。转基因烟草的Northern杂交检测显示,30%PEG诱导条件下CBF2基因在诱导30 min之后持续较高的表达水平,且表达水平受胁迫诱导程度的影响。抗旱生理指标测定显示,干旱条件下转基因烟草植株的脯氨酸含量迅速增高,远超于野生型;而丙二醛含量的增长幅度要比野生型植株小很多,且随着胁迫时间的延长,转基因植株体内的丙二醛含量逐步趋于稳定。试验证明,CBF2基因在提高农作物的抗旱性方面具有极大的利用价值。     

Overexpression of osmotin gene confers tolerance to salt and drought stresses in transgenic tomato (Solanum lycopersicum L.)

Abiotic stresses, especially salinity and drought, are major limiting factors for plant growth and crop productivity. In an attempt to develop salt and drought tolerant tomato, a DNA cassette containing tobacco osmotin gene driven by a cauliflower mosaic virus 35S promoter was transferred to tomato (Solanum lycopersicum) via Agrobacterium-mediated transformation. Putative T0 transgenic plants were screened by PCR analysis. The selected transformants were evaluated for salt and drought stress tolerance by physiological analysis at T1 and T2 generations. Integration of the osmotin gene in transgenic T1 plants was verified by Southern blot hybridization. Transgenic expression of the osmotin gene was verified by RT-PCR and northern blotting in T1 plants. T1 progenies from both transformed and untransformed plants were tested for salt and drought tolerance by subjecting them to different levels of NaCl stress and by withholding water supply, respectively. Results from different physiological tests demonstrated enhanced tolerance to salt and drought stresses in transgenic plants harboring the osmotin gene as compared to the wild-type plants. The transgenic lines showed significantly higher relative water content, chlorophyll content, proline content, and leaf expansion than the wild-type plants under stress conditions. The present investigation clearly shows that overexpression of osmotin gene enhances salt and drought stress tolerance in transgenic tomato plants.     

The Arabidopsis GIBBERELLIN METHYL TRANSFERASE 1 suppresses gibberellin activity,reduces whole‐plant transpiration and promotes drought tolerance in transgenic tomato

Previous studies have shown that reduced gibberellin (GA) level or signal promotes plant tolerance to environmental stresses, including drought, but the underlying mechanism is not yet clear. Here we studied the effects of reduced levels of active GAs on tomato (Solanum lycopersicum) plant tolerance to drought as well as the mechanism responsible for these effects. To reduce the levels of active GAs, we generated transgenic tomato overexpressing the Arabidopsis thaliana GA METHYL TRANSFERASE 1 (AtGAMT1) gene. AtGAMT1 encodes an enzyme that catalyses the methylation of active GAs to generate inactive GA methyl esters. Tomato plants overexpressing AtGAMT1 exhibited typical GA‐deficiency phenotypes and increased tolerance to drought stress. GA application to the transgenic plants restored normal growth and sensitivity to drought. The transgenic plants maintained high leaf water status under drought conditions, because of reduced whole‐plant transpiration. The reduced transpiration can be attributed to reduced stomatal conductance. GAMT1 overexpression inhibited the expansion of leaf‐epidermal cells, leading to the formation of smaller stomata with reduced stomatal pores. It is possible that under drought conditions, plants with reduced GA activity and therefore, reduced transpiration, will suffer less from leaf desiccation, thereby maintaining higher capabilities and recovery rates.     

Effect of Elevated CO2 and Drought on Soil Microbial Communities Associated with Andropogon gerardii

Our understanding of the effects of elevated atmospheric CO2, singly and In combination with other environmental changes,on plant-soil interactions is incomplete. Elevated CO2 effects on C4 plants, though smaller than on C3 species, are mediated mostly via decreased stomatal conductance and thus water loss. Therefore, we characterized the interactive effect of elevated CO2 and drought on soil microbial communities associated with a dominant C4 prairie grass, Andropogon gerardii Vitman. Elevated CO2 and drought both affected resources available to the soil microbial community. For example, elevated CO2 increased the soil C:N ratio and water content during drought, whereas drought alone decreased both. Drought significantly decreased soil microbial biomass. In contrast, elevated COz increased biomass while ameliorating biomass decreases that were induced under drought. Total and active direct bacterial counts and carbon substrate use (overall use and number of used sources) increased significantly under elevated CO2. Denaturing gradient gel electrophoresis analysis revealed that drought and elevated CO2, singly and combined, did not affect the soil bacteria community structure.We conclude that elevated CO2 alone increased bacterial abundance and microbial activity and carbon use, probably in response to increased root exudation. Elevated CO2 also limited drought-related impacts on microbial activity and biomass,which likely resulted from decreased plant water use under elevated CO2. These are among the first results showing that elevated CO2 and drought work in opposition to modulate plant-associated soil-bacteria responses,which should then Influence soil resources and plant and ecosystem function.     

Interactive Effects of Drought Stresses and Elevated CO2 Concentration on Photochemistry Efficiency of Cucumber Seedlings

To reveal and quantify the interactive effects of drought stresses and elevated CO2 concentration [CO2] on photochemistry efficiency of cucumber seedlings, the portable chlorophyll meter was used to measure the chlorophyll content, and the Imaging-PAM was used to image the chlorophyll fluorescence parameters and rapid light response curves (RLC) of leaves in two adjacent greenhouses. The results showed that chlorophyll content of leaves was reduced significantly with drought stress aggravated. Minimal fluorescence (Fo) was increased while maximal quantum yield of PSII (Fv/Fm) decreased significantly by severe drought stress. The significant decrease of effective quantum yield of PSll (Y(Ⅱ)) accompanied by the significant increase of quantum yield of regulated energy dissipation (Y(NPQ)) was observed under severe drought stress condition, but there was no change of quantum yield of nonregulated energy dissipation (Y(NO)). We detected that the coefficient of photochemical quenching (Qp) decreased, and non-photochemical quenching (NPQ) increased significantly under severe drought stress. Furthermore, we found that maximum apparent electron transport rate (ETRmax) and saturating photosynthetically active radiation (PPFDsat) decreased significantly with drought stress aggravated. However, elevated [CO2] significantly increased FvlFm, Qp and PPFDsat, and decreased NPQ under all water conditions, although there were no significant effects on chlorophyll content, Fo, Y(Ⅱ), Y(NPQ), Y(NO) and ETRmax. Therefore, it is concluded that CO2-fertilized greenhouses or elevated atmospheric [CO2] in the future could be favorable for cucumber growth and development, and beneficial to alleviate the negative effects of drought stresses to a certain extent.     

Constitutive expression of CaXTH3, a hot pepper xyloglucan endotransglucosylase/hydrolase, enhanced tolerance to salt and drought stresses without phenotypic defects in tomato plants (Solanum lycopersicum cv. Dotaerang)

The hot pepper xyloglucan endo-trans-gluco-sylase/hydrolase (CaXTH3) gene that was inducible by a broad spectrum of abiotic stresses in hot pepper has been reported to enhance tolerance to drought and high salinity in transgenic Arabidopsis. To assess whether CaXTH3 is a practically useful target gene for improving the stress tolerance of crop plants, we ectopically over-expressed the full-length CaXTH3 cDNA in tomato (Solanum lycopersicum cv. Dotaerang) and found that the 35S:CaXTH3 transgenic tomato plants exhibited a markedly increased tolerance to salt and drought stresses. Transgenic tomato plants exposed to a salt stress of 100?mM NaCl retained the chlorophyll in their leaves and showed normal root elongation. They also remained green and unwithered following exposure to 2?weeks of dehydration. A high proportion of stomatal closures in 35S:CaXTH3 was likely to be conferred by increased cell-wall remodeling activity of CaXTH3 in guard cell, which may reduce transpirational water loss in response to dehydration stress. Despite this increased stress tolerance, the transgenic tomato plants showed no detectable phenotype defects, such as abnormal morphology and growth retardation, under normal growth conditions. These results raise the possibility that CaXTH3 gene is appropriate for application in genetic engineering strategies aimed at improving abiotic stress tolerance in agriculturally and economically valuable crop plants.     

拟南芥CBF1与植物对低温和干旱的抗性

对冷驯化过程中基因表达差异的认识,使抗冻基因(COR)的克隆及其功能的分析成为研究冷驯化过程的主要目标。在拟南芥和其他抗冻植物中,分离了许多COR基因,这些基因对植物抗冻起着非常重要的作用。在拟南芥COR调控的研究中,发现了CBF转录因子的基因家族,其中CBF1能调控一组COR基因的表达。近年来,在冷敏植物如番茄和玉米中也发现了CBF类似基因,拟南芥CBF1基因在转基因番茄中的过量表达提高了植株的抗寒和抗旱性。这一研究结果展示了拟南芥CBF1类似基因的应用可能为冷敏植物抗寒和抗旱性的品种改良提供一条新的途径。