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为了探究 GbCDPK83基因在海岛棉响应干旱胁迫中的功能。利用PCR技术克隆 GbCDPK83基因,采用生物信息学方法分析GbCDPK83蛋白的理化性质、结构特征和在细胞中的位置,通过基因重组技术构建VIGS沉默载体并侵染棉花。本研究成功构建了沉默表达载体,沉默植株中 GbCDPK83的表达明显被抑制。干旱胁迫后, GbCDPK83沉默植株叶片比对照萎蔫更严重,相对含水量显著降低,相对电导率和丙二醛含量显著上升,脯氨酸含量升高但低于空载体及非转基因植株。沉默 GbCDPK83使海岛棉耐旱性减弱。  相似文献   

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棉花干旱诱导MYB类转录因子GhRAX3的功能分析   总被引:2,自引:0,他引:2  
【目的】挖掘、分析响应干旱诱导的MYB转录因子的功能,为棉花抗旱研究和育种提供参考。【方法】以已知受干旱诱导表达的GbMYB5为检索项,Blastx检索NCBI的非冗余蛋白nr数据库中与GbMYB5具有相似序列的棉花MYB转录因子,通过荧光定量PCR验证获得干旱应答相关的MYB转录因子GhRAX3。将GhRAX3-GFP融合蛋白表达载体通过农杆菌注射法在烟草叶片瞬时表达,观察亚细胞定位荧光信号。将棉花曲叶病毒CLCrV介导的基因沉默载体CLCrV﹕GhRAX3通过农杆菌浸润法注射棉花子叶,荧光定量PCR检测棉花GhRAX3表达水平,对GhRAX3沉默的棉花植株进行干旱胁迫处理,观察表型变化,并检测其叶片失水率、叶片含水量、总抗氧化物酶活性、丙二醛含量和离子渗透率等抗旱相关生理指标。【结果】Blastx检索结果发现,与GbMYB5相比,GhRAX3覆盖率达38%,与GbMYB5有79%的相似性,编码一个R2R3-MYB转录因子。GhRAX3响应干旱诱变表达,在18%(v/v)PEG 6000诱导处理0.5 h后即显著上调表达5倍,在48 h后达上调表达33倍。亚细胞定位结果显示GhRAX3-GFP4融合蛋白仅在细胞核有明显的绿色荧光信号,表明GhRAX3定位在细胞核中。CLCrV病毒诱导GhRAX3沉默后,GhRAX3在沉默株的表达量仅为野生型的41%,表明GhRAX3表达已被抑制。取干旱处理前的棉花植株同部位的叶片,测定单位重量叶片在0-7 h内的失水量,发现GhRAX3沉默植株的叶片失水率显著高于野生型和空载体对照植株。在18%(v/v)PEG 6000水溶液处理24 h或在自然干旱15 d后,GhRAX3沉默的棉花植株萎蔫程度比野生型和空载体对照植株严重。自然干旱处理7 d后,检测叶片含水量、总抗氧化物酶活性、丙二醛含量和离子渗透率等抗旱相关生理指标,发现GhRAX3沉默植株的叶片相对含水量为82%,总抗氧化物酶活性为1.29 U·mg-1,而野生型和空载体对照植株的叶片相对含水量则分别为89%和91%,总抗氧化物酶活性分别为3.44和3.19 U·mg-1,GhRAX3沉默植株的叶片相对含水量和总抗氧化物酶活性都显著低于野生型和空载体对照植株;相反,GhRAX3沉默植株叶片的离子渗漏率为78.54%,丙二醛含量为74.20 nmol·mg-1 FW,而野生型和空载体对照植株叶片的离子渗漏率分别为44.98%和47.45%,丙二醛含量分别为44.90和47.29 nmol·mg-1 FW,GhRAX3沉默植株叶片的离子渗漏率和丙二醛含量均显著高于野生型和空载体对照。【结论】GhRAX3响应干旱胁迫诱导,抑制GhRAX3表达致使棉花在干旱胁迫条件下的叶片相对含水量和总抗氧化物酶活性显著降低,叶片离子渗漏率和丙二醛含量显著升高,使棉花耐干旱胁迫能力下降。  相似文献   

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Relationships between plant water status and gas exchange parameters at increasing levels of water stress were determined in Algerie loquats which grown in 50 I pots. Changes in soil water content and stem water potential and their effects on stomatal conductance(G_s) and net photosynthesis(P_n) rate were followed in control plants and in plants without irrigation until the latter reached near permanent wilting point and some leaf abscission took place. Then, the irrigation was restarted and the comparison repeated. Soil water content and stem water potential gradually diminished in response to drought reaching the minimum values of 0.9 mm and –5.0 MPa, respectively, 9 days after watering suspension. Compromised plant water status had drastic effects on G_s values that dropped by 97% in the last day of the drought period. P_n was diminished by 80% at the end of the drought period. The increasing levels of water stress did not cause a steady increase in leaf temperature in non-irrigated plants. Non-irrigated plants wilted and lost some leaves due to the severity of the water stress. However, all non-irrigated plants survived and reached similar P_n than control plants just a week after the irrigation was restarted, confirming drought tolerance of loquat and suggesting that photosynthesis machinery remained intact.  相似文献   

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Maize is one of the most important crops worldwide, but it suffers from salt stress when grown in saline-alkaline soil. There is therefore an urgent need to improve maize salt tolerance and crop yield. In this study, the SsNHX1 gene of Suaeda salsa, which encodes a vacuolar membrane Na+/H+ antiporter, was transformed into the maize inbred line 18-599 by Agrobacterium-mediated transformation. Transgenic maize plants overexpressing the SsNHX1 gene showed less growth retardation when treated with an increasing NaCl gradient of up to 1%, indicating enhanced salt tolerance. The improved salt tolerance of transgenic plants was also demonstrated by a significantly elevated seed germination rate (79%) and a reduction in seminal root length inhibition. Moreover, transgenic plants under salt stress exhibited less physiological damage. SsNHX1-overexpressing transgenic maize accumulated more Na+ and K+ than wild-type (WT) plants particularly in the leaves, resulting in a higher ratio of K+/Na+ in the leaves under salt stress. This result revealed that the improved salt tolerance of SsNHX1-overexpressing transgenic maize plants was likely attributed to SsNHX1-mediated localization of Na+ to vacuoles and subsequent maintenance of the cytosolic ionic balance. In addition, SsNHX1 overexpression also improved the drought tolerance of the transgenic maize plants, as rehydrated transgenic plants were restored to normal growth while WT plants did not grow normally after dehydration treatment. Therefore, based on our engineering approach, SsNHX1 represents a promising candidate gene for improving the salt and drought tolerance of maize and other crops.  相似文献   

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SLB9基因是BES1转录因子家族中的一员,BES1在调控油菜素内酯(BR)基因响应中发挥关键作用。研究证明BES1基因受多种胁迫诱导,介导BR信号,调节植物耐旱等抗逆性。采用基因沉默(VIGS)技术,通过农杆菌介导法构建SLB9基因沉默植株,干旱胁迫处理沉默植株,通过观察沉默植株与对照植株表型差异及生理生化指标变化,研究SLB9基因表达变化对番茄抗旱性的影响。结果表明,SLB9基因沉默植株抗旱性明显低于对照植株,推测SLB9基因下调表达降低番茄抗旱性。  相似文献   

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前期研究已完成番茄WRKY转录因子家族分析,发现番茄WRKYⅡa和Ⅱb亚族基因多与抗逆相关,因此文章选取6个WRKYⅡa和Ⅱb亚族基因Sl WRKY13、Sl WRKY24、Sl WRKY31、Sl WRKY50、Sl WRKY62、Sl WRKY63,运用q RT-PCR分析方法,分析逆境胁迫下表达模式。结果表明,Sl WRKY24干旱、盐、低温胁迫下表达受抑制,另外5个基因在三种胁迫处理中,除干旱胁迫处理Sl WRKY50基因表达量下降,其余均不同程度上调表达。低温胁迫处理下Sl WRKY50基因表达量与对照相比表现极显著差异。Sl WRKY50基因沉默分析结果表明,Sl WRKY50基因沉默后,下调Sl WRKY50基因表达,干旱、高盐胁迫下植株叶片Pro、SOD、MAD含量水平与对照相比变化较小,低温胁迫下番茄植株叶片Pro和SOD含量低于对照,MAD提高含量,植株对低温逆境胁迫耐受能力下降。研究为进一步探讨WRKY基因家族功能提供参考及理论依据。  相似文献   

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Alfalfa (Medicago sativa L.) is an important forage crop in the world and it is of great significance for the improvement of its salt tolerance. To improve salt tolerance in alfalfa, a rice ascorbate peroxidase gene (OsAPX2) was introduced into alfalfa using Agrobacterium tumefaciens-mediated transformation with marker gene bar. The different T-DNA insertions in T1 transgenic alfalfa were identified by Southern hybridization. Three independent T2 transgenic lines were selected for stress analysis and the results showed that all of them were salt tolerant compared with wild-type plants. The transgenic plants had low levels of H2O2, malondialdehyde and relative electrical conductivity under salt and drought stresses. Moreover, the contents of chlorophyll and proline, and APX activity were high in transgenic plants under salt and drought stresses. Taken together, the overexpression of OsAPX2 enhances salt tolerance in alfalfa through scavenging reactive oxygen species.  相似文献   

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Grapevine growing areas are increasingly affected by drought, which has greatly limited global wine production and quality. DEAD-box is one of the largest subfamilies of the RNA helicase family, and its members play key roles in the growth and development of plants and their stress responses. Previous studies have shown the potential of DEAD-box genes in the drought stress responses of Arabidopsis and tomato, rice, and other crop species. However, information about DEAD-box genes in grapevine remains limited. In this report, a total of 40 DEAD-box genes were identified in grapevine and their protein sequence characteristics and gene structures were analyzed. By comparing the expression profiles of VviDEADRHs in response to drought stress in different grapevine varieties, nine candidate genes (VviDEADRH10c, -13, -22, -25a, -25b, -33, -34, -36, and -39) were screened based on expression profiling data. Combined with qRT-PCR results, VviDEADRH25a was selected for functional verification. Heterologous overexpression of VviDEADRH25a in Arabidopsis showed the transgenic plants were more sensitive to drought stress than the control. Both electrolyte permeability and malondialdehyde content were significantly increased in transgenic plants, whereas the chlorophyll content and superoxide dismutase (SOD), peroxidase (POD), catalase (CAT), and ascorbate peroxidase (APX) enzyme activities were significantly decreased. Furthermore, VviDEADRH25a-overexpressing plants showed down-regulated expression levels of several drought stress-related marker genes, namely AtCOR15a, AtRD29A, AtERD15, and AtP5CS1, which indicated that they participated in the drought stress response. In summary, this study provides new insights into the structure, evolution, and participation of DEAD-box RNA helicase genes in the response to drought stress in grapevines.  相似文献   

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Trehalose plays an important role in metabolic regulation and abiotic stress tolerance in a variety of organisms. In plants, its biosynthesis is catalyzed by two key enzymes: trehalose-6-phosphate synthase(TPS) and trehalose-6-phosphate phosphatase(TPP). In the present study, a TPS gene, named IbTPS, was first isolated from sweetpotato(Ipomoea batatas(L.) Lam.) cv. Lushu 3 by rapid amplification of cDNA ends(RACE). The open reading frame(ORF) contained 2 580 nucleotides encoding 859 amino acids with a molecular weight of 97.433 kDa and an isoelectric point(pI) of 5.7. The deduced amino acid sequence showed high identities with TPS of other plants. Real-time quantitative PCR analysis revealed that the expression level of IbTPS gene was significantly higher in stems of Lushu 3 than in its leaves and roots. Subcellular localization analysis in onion epidermal cells indicated that IbTPS gene was located in the nucleus. Transgenic tobacco(cv. Wisconsin 38) plants over-expressing IbTPS gene exhibited significantly higher salt tolerance compared with the control plant. Trehalose and proline content was found to be significantly more accumulated in transgenic tobacco plants than in the wild-type and several stress tolerance related genes were up-regulated. These results suggest that IbTPS gene may enhance salt tolerance of plants by increasing the amount of treahalose and proline and regulating the expression of stress tolerance related genes.  相似文献   

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为深入研究褪黑素的分子生物学功能,根据番茄基因组数据库的SlSNAT基因序列信息设计引物,以耐盐番茄材LA1401(PI365967)叶片RNA反转录得到的cDNA为模板,利用高保真酶克隆了番茄的褪黑素合成酶基因SlSNAT,基因CDS(coding sequence)序列全长为768 bp,共编码255个氨基酸。利用酶切连接的方法,构建了该基因的超表达载体。实时定量PCR结果表明,SlSNAT基因在叶片中的表达量最高,显著高于其在花、果实、根、种子和萼片中的表达量。在不同非生物逆境处理下的表达结果显示,在干旱处理条件下的表达量最高,其次是在甘露醇的渗透胁迫逆境,在这2种胁迫条件下的表达量均显著高于其在过氧化氢、氯化钠、低温和褪黑素诱导下的表达量。另外,在盐胁迫条件下,SlSNAT基因在根部的表达量受到明显抑制。  相似文献   

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Based on the DNA sequence of ACS9, two produced fragments were subcloned into binary vector pCAMBIA1300 in antisense and sense orientations, and the generated RNA interference (RNAi) vector was then transformed into Arabidopsis thaliana. The stress resistance function of ACS9 gene in Arabidopsis thaliana was researched by determination of stress resistance physiologic indexes, NaCl and PEG6000 resistance. The results showed that the inhibition of ACS9 expression enhanced the sensitivity to high concentration NaCl (150 mmol/L) and PEG6000 (7%) in Arabidopsis thaliana seeding stage. The proline contents and water loss rates in transgenic plants were 0.68 and 1.4 times higher than those in the wild-type leaves, respectively, indicating that the inhibition of ACS9 expression due to salt and drought resistant was reduced and suggested that ACS9 gene played important roles in plant salt and drought tolerance.  相似文献   

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外源褪黑素对干旱胁迫下番茄叶片光合作用的影响   总被引:1,自引:0,他引:1  
杨小龙  须晖  李天来  王蕊 《中国农业科学》2017,50(16):3186-3195
【目的】褪黑素是一种广泛存在于高等植物体内的小分子物质,被认为是一种新的植物生长调节剂和生物刺激剂,对于提高植物抗逆性具有重要作用。探索外源褪黑素对干旱胁迫下番茄叶片光合作用的影响,为揭示褪黑素调节植物抗逆性的机制打下基础。【方法】以番茄‘辽园多丽’为试材,首先采用叶片喷施和根施不同浓度褪黑素进行预处理:CK:叶片喷施清水、根施50 m L清水;R5、R50、R100、R150、R250:叶片喷清水,分别根施50m L 5、50、100、150和250μmol·L~(-1)褪黑素;L5、L50、L100、L150、L250:根施50 m L清水,叶片分别喷施5、50、100、150和250μmol·L~(-1)褪黑素;连续处理3 d后将植株移至温室中,以不浇水作为干旱处理(其中CK0:叶片喷施清水、根施50 m L清水预处理后正常浇水,CK1:叶片喷施清水、根施50 m L清水预处理后干旱处理)。干旱胁迫5 d后,通过比较暗适应下PSII最大光化学量子产量Fv/Fm和PSI最大氧化状态Pm,确定根施和叶片喷施的最佳浓度处理。然后利用光合荧光同步测量系统分析根施和叶片喷施褪黑素对干旱胁迫下番茄叶片气体交换参数,PSII和PSI的光能分配和电子传递速率,类囊体膜的完整性和ATP酶活性的调节。【结果】根施和叶片喷施不同浓度褪黑素均提高了干旱胁迫下番茄叶片的Fv/Fm和Pm,并且随浓度增加表现出先升高后降低的趋势,L100和R100处理下的Fv/Fm和Pm最大,显著高于对照。L100和R100显著缓解了干旱胁迫对气体交换参数的抑制,其中叶片净光合速率(Pn)分别为2.04和1.71μmol·m~(-2)·s~(-1),显著高于对照(CK1)(0.52μmol·m~(-2)·s~(-1));蒸腾速率(E)分别为0.66和0.54 mmol·m~(-2)·s~(-1),显著高于CK1(0.25 mmol·m~(-2)·s~(-1)),并且显著提高了番茄叶片气孔导度(GH2O)和最大水分利用效率(WUE),降低了气孔限制值(Ls),而L100优于R100处理。快速光响应曲线结果表明L100和R100处理提高了干旱胁迫下番茄幼苗PSII的光化学反应效率Fv'/Fm'及PSII光化学淬灭系数q P,表明褪黑素处理更利于干旱胁迫下番茄叶片PSII光化学反应的高效进行;干旱胁迫下番茄幼苗环式电子传递速率得到显著加强,而L100和R100处理降低了环式电子传递速率,但加强了线性电子传递速率,且L100处理下番茄叶片ETRI和ETRII均高于R100处理;L100、R100处理提高了干旱胁迫下番茄叶片的Y(I)、Y(II),表明褪黑素处理有利于干旱胁迫下番茄叶片吸收光能向光化学反应的方向分配;暗适应后,L100和R100处理番茄叶片P515诱导曲线均高于CK1,照光后,CK0处理番茄幼苗P515信号快速下降,其次是L100和R100处理,而CK1处理降低较慢,表明褪黑素具有保护叶绿体类囊体膜和ATP合成酶免受干旱胁迫伤害的作用。【结论】根施和叶片喷施外源褪黑素能缓解干旱胁迫对番茄幼苗光合性能的抑制,加强光合运转效率,而叶片喷施是一种更简单高效的处理方式;褪黑素能加强作物光合作用对环境胁迫的适应性,对于农作物的生长发育具有调节作用。  相似文献   

17.
【目的】分析棉花中钠尿肽GhPNP1的结构特征、表达模式以及耐旱功能,并分析其耐旱机制,为将该基因应用于作物改良奠定基础。【方法】通过对从植物中水平转移到大丽轮枝菌中的钠尿肽基因AVE1进行同源性搜索,得到与AVE1蛋白序列相似度较高的其他物种的蛋白序列;使用MEGA5软件对AVE1蛋白序列及其同源序列进行多序列比对分析并构建同源物种间系统进化树;利用MEGA和expasy在线工具进行蛋白序列分析;并根据编码该蛋白的核酸序列设计引物在陆地棉品种奥3503中克隆到其同源基因GhPNP1。使用多种生物信息学软件分析GhPNP1的分子特性,包括GhPNP1编码蛋白的等电点、分子量、信号肽、进化关系等进行预测分析。利用实时荧光定量PCR(qRT-PCR)分析GhPNP1在不同器官部位的组织表达模式以及受到PEG模拟干旱胁迫处理后的表达模式。将GhPNP1的cDNA序列连入CLCrV沉默载体中,构建GhPNP1的病毒诱导基因沉默载体CLCrV:GhPNP1,转入农杆菌,并通过和辅助载体CLCrVB共侵润2叶期幼苗进行叶片注射,获得GhPNP1的沉默植株。利用PEG模拟干旱处理沉默植株检测其耐旱性,并测定沉默植株的失水率、相对含水量、丙二醛(MDA)含量、总抗氧化活性(T-AOC水平)、离子渗漏率等与植物抗逆相关的生理指标。【结果】从陆地棉奥3503中克隆到的GhPNP1的开放阅读框长度为396 bp,编码131个氨基酸,信号肽长度为15个氨基酸,通过系统进化树分析GhPNP1编码的蛋白含有保守的钠尿肽结构域,与可可树的PNP蛋白进化关系最近。GhPNP1在棉花植株的根、茎、叶中均表达且在茎中表达量较高,PEG模拟干旱处理后根、茎、叶中的GhPNP1均上调表达。GhPNP1沉默后棉花植株耐旱性显著降低。在干旱条件下,GhPNP1沉默植株的MDA含量、离子渗漏率、叶片失水率均高于对照的未沉默植株;而沉默植株的总抗氧化能力(T-AOC水平)、相对含水量显著低于对照的未沉默植株。【结论】从棉花中克隆得到一个植物钠尿肽基因,受干旱胁迫诱导上调表达,沉默后耐旱性降低。推测GhPNP1可能通过cGMP信号途径参与棉花的干旱胁迫,在干旱胁迫下对棉花耐旱性起正调控作用。  相似文献   

18.
Silicon can improve drought tolerance of plants,but the mechanism still remains unclear.Previous studies have mainly concentrated on silicon-accumulating plants,whereas less work has been conducted in silicon-excluding plants,such as tomato(Solanum lycopersicum L.).In this study,we investigated the effects of exogenous silicon(2.5 mmol L~(–1))on the chlorophyll fluorescence and expression of photosynthesis-related genes in tomato seedlings(Zhongza 9)under water stress induced by 10%(w/v)polyethylene glycol(PEG-6000).The results showed that under water stress,the growth of shoot and root was inhibited,and the chlorophyll and carotenoid concentrations were decreased,while silicon addition improved the plant growth and increased the concentrations of chlorophyll and carotenoid.Under water sterss,chlorophyll fluorescence parameters such as PSII maximum photochemical efficiency(F_v/F_m),effective quantum efficiency,actual photochemical quantum efficiency(Ф_(PSII)),photosynthetic electron transport rate(ETR),and photochemical quenching coefficient(q_P)were decreased;while these changes were reversed in the presence of added silicon.The expressions of some photosynthesis-related genes including PetE,PetF,PsbP,PsbQ,PsbW,and Psb28 were down-regulated under water stress,and exogenous Si could partially up-regulate their expressions.These results suggest that silicon plays a role in the alleviation of water stress by modulating some photosynthesis-related genes and regulating the photochemical process,and thus promoting photosynthesis.  相似文献   

19.
采用温室有机土盆栽试验研究了不同盐浓度(N aC l浓度分别为5和10 g/L)胁迫下,丛枝菌根真菌(AM F)对番茄氧自由基(O2.-)产生速率,丙二醛(M DA)含量,细胞膜透性,谷胱甘肽过氧化物酶(G SH-Px)活性和叶片相对含水量(RW C)等生理指标的影响。结果表明,在盐胁迫下,接种和未接种AM F的番茄均随盐浓度的增加和盐胁迫时间的延长,叶片和根系中M DA含量、叶片中O2.-产生速率和细胞膜透性持续增加,RW C下降;与未接种番茄相比,接种AM F能显著减少番茄植株叶片中M DA的积累,降低O2.-产生速率和细胞膜透性,增加RW C,因而减缓了盐胁迫对番茄细胞膜的伤害,增强了番茄的耐盐性。盐胁迫下,接种AM F番茄的G SH-Px活性显著高于未接种株,可见接种AM F增强了番茄叶片的G SH-Px活性,减轻了活性氧对植株细胞膜的伤害,提高了番茄的耐盐性。  相似文献   

20.
Introduction of the FeSOD gene enhanced the stability of the photosynthetic apparatus of plants to the action of oxidative stress caused by UV irradiation. The expression of Arabidopsis thaliana FeSOD gene, targeting the enzyme in chloroplasts due to a signal sequence, leaded to significant changes in ultrastructure of cell subcompartments of tobacco and tomato leaves. The activity of superoxide dismutase in leaves of transgenic tomato plants exceeded the value of activity of this enzyme of control plants. Transgenic tobacco plants showed increasing in SOD activity compared with control non-transgenic tobacco. The activity of AP in the leaves of transgenic tobacco and tomato plants was similar with that of control non-transgenic plants, but activity of one accession of transgenic tomato, which is also characterized by high values of SOD activity, exceeded the value of control plant. Differences in ultrastructural organization of chloroplasts in the cells of transgenic and control tobacco and tomato plants have been manifested in a strong enlargement in the size of plastoglobuli. These distinctions were evident especially in the cells of the leaf parenchyma of transgenic tomato as well as transgenic tobacco. Also, a quantity of starch grains in the plastids of guard cells was increased. Chloroplasts in the cells of leaf parenchyma in transgenic plants contained less a starch grains than in wild-type plants.  相似文献   

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