RNAi沉默烟草GA 20-氧化酶基因研究

赤霉素是一种高效能的广谱植物生长调节剂，为五大植物激素之一，具有重要的生物学功能。目前利用赤霉素突变体研究生物合成途径和信号转导已经成为热点。 GA 20-氧化酶是GA生物合成中的一类关键酶，它位于GA合成途径的中心位置。本研究根据烟草（Nicotiana tabacum）GA 20-氧化酶基因序列，设计2对分别含有特定酶切位点的特异引物，以烟草基因组DNA为模板，扩增目的基因(约250 bp)片段。将正、反向目的片段分别插入中间载体的内含子两侧，再经BamH I和Sac I双酶切回收约700 bp的目的片段，插入到双元载体质粒p2355中，成功构建了含GA 20-氧化酶基因片段反向重复序列的植物表达载体p23700。分别将p2355质粒和p23700质粒导入根癌农杆菌（Agrobacterium tumefaciens）EHA105中并转化烟草叶片细胞，经卡那霉素选择培养，PCR及GUS组织染色鉴定，获得转基因烟草植株。以EHA105-p2355转化的烟草，获得41株转基因植株，均没有矮化表型；而以EHA105-p23700转化的烟草，获得转基因植株14株，其中具有矮化表型的烟草10株，表明反向重复序列转录产物能形成发夹RNA(hpRNA)，产生小分子干扰RNA（small interferring RNA，简称siRNA），干扰目的基因的表达。 赤霉素含量测定表明矮化植株中赤霉素合成途径的最终产物GA3总含量明显低于野生型烟草植株。荧光定量PCR结果表明，矮化转基因烟草的GA 20-氧化酶基因表达量受到明显抑制，表达量明显低于野生型对照。同时对上游内根-贝壳杉合成酶（Ent-kaurene synthase，KS）基因，下游的GA-3β羟化酶基因进行了RT-PCR分析，结果显示上游基因的表达没有规律性变化，而下游基因表达量亦降低。上述结果表明，GA 20-氧化酶基因的表达被有效地干扰了，表达受到抑制，从而影响植株体内GA3的合成，影响植株的生长发育，导致植株矮化。并推测，GA 20-氧化酶基因受到抑制，可能影响下游基因的表达。并且通过干旱胁迫测试，发现矮化植株相对于野生型植株及不含干扰片段的转基因植株，对干旱的耐受力有了很大的提高，具有更强的耐受力。 研究结果为进一步进行相关研究奠定基础。 Gibberellin(GA) is an efficient plant growth regulator. As one of five major plant hormones, it plays an important biological function. Using GA mutant for investigating biosynthetic pathways and signal transduction has become high lights. GA 20-oxidase is a crucial enzyme involved in gibberellin biosynthesis. According to tobacco (Nicotiana tabacum) GA 20-oxidase enzyme gene sequence and based on binary vector p2355, we constructed a plant expression vector p23700, which habors an inverted repeat DNA fragment of GA 20-oxidase gene drivered by Cauliflower mosaic virus promtor (CaMV 35Sp). Binary plasmid p2355 had no inverted repeat DNA fragment of GA 20-oxidase gene. The vector p2355 and p23700 were introduced into Agrobacterium tumefaciens EHA105 and tobacco leaf transformation was conducted. After selected by kanamycin and characterized by PCR and GUS hischemical reaction, transsgenic plants were obtained. Fourtheen transgenic plants, which were transformed by EHA105-p23700, were obtained. Among them, 10 were dwarf mutants. However, 41 transgenic plants with the same normal phenotype as wild type,which were transformed by EHA105-p2355, were obtained. Analysis of Gibberellin contents showed that it was lower in dwarf mutants than in normal phenotype plants. Moreover, comparing to normal phenotype plants including wild type and transgenic plants with no interference fragment, the drought tolerance of dwarf plants have greatly increased. And their proline content increased obviously after drought test. Fluorescence quantitative real time PCR (RT-PCR) showed that GA 20-oxidase gene expression was significantly inhibited in dwarf transgenic tobacco. Meanwhile, the expression of the upstream gene ent-kaurene synthase (KS) gene and downstream gene GA-3β hydroxylase gene was also detected by RT-PCR. The results presented that KS gene expression had no regular change while GA-3β hydroxylase gene expression reduced. It implied that inhibiting GA 20-oxidase gene probably reduce the expression of downstream genes. The results showed that the transcriptional products of the foreign inverted repeat fragment can form hairpin RNA (hpRNA) to induce RNAi. It presented that GA 20-oxidase gene expression was effectively interfered, resulting in reducing GA3 synthesis and inhibiting plant growth and development, then dwarf plants were produced. However, the dwarf plants had higher tolerance of drought.

厌氧好氧一体化生物反应器处理发酵废水的中试研究

一体化反应器由于投资少、占地小、管理运行方便等优点而备受青睐。但现有的一体化反应器大都适用于处理中低浓度废水，耐受负荷普遍偏低。本课题研制出新型高效的厌氧好氧一体化生物反应器，旨在通过反应器结构优化、高效微生物载体研制、配合高效微生物菌剂技术处理中高浓度有机废水，实现高效和低耗，降低设备造价，提高反应器运行稳定性。 首先开展了菌剂对废水的适配试验。采用15种不同的微生物菌剂，以葡萄糖配水、中药提取废水、啤酒废水、氨氮配水为基质，分别测定了微生物菌剂的耗氧速率和厌氧比产甲烷速率，以其为指标比较了各菌剂对废水的适配性。根据结果选择活性高的14＃、8＃、10＃菌剂，在试验室进行了菌剂对废水的连续处理试验，取得良好的处理效果，为菌剂在厌氧好氧一体化生物反应器的小试、中试中的应用奠定了基础。 经小试研究后，又对厌氧好氧一体化生物反应器进行了处理发酵废水的中试研究。试验结果表明，反应器启动快，系统有机负荷2.72 kgCODm-3d-1时整个反应器去除率保持在84.5％～93.19％，在30多天内一次启动成功。冲击负荷试验中，系统总有机负荷最高可达到8.88 kgCODm-3d-1，系统去除率稳定在88.10％～96.88%，说明反应器处理效率高，抗冲击能力强。稳定运行期间，COD去除率可达90％以上，各项指标都能达到国家排放标准。 此外，对反应器配套系统高效菌剂、高分子复合颗粒载体进行了研究。结果显示，菌剂与反应器适配良好，各功能区形成了丰富、高活性的微生物，厌氧区颗粒污泥TS高达83.9 gL-1，VS/TS为56.9％～57.4％，比产甲烷活性为280～350 mLCH4 gvss-1d-1；好氧区固定化微生物TS高达1.921 gL-1，VS/TS为94.02～94.30％。对载体性能的研究表明，此高分子复合颗粒载体密度适中，易于流化，不易流失；粗糙多空，易于挂膜；且无生物毒害作用，稳定安全，是一种优良的生物载体。反应器各功能区对废水的降解过程分析，说明了反应器、菌剂、载体适配良好，在其协同作用下，实现了污染物的高效降解。 The integrated reactors were popular because of their characteristics such as little investment, small occupation of land, convenient of manage and running etc. But the present integrated reactors were mostly applied for treating wastewater of low concentration, the load tolerance was generally on the low side. A new type integrated anaerobic-aerobic bio-reactor was developed, which was conducted to treating organic wastewater of middle or high concentration by optimization of reactor structure, development of efficient microbe carrier and adaptation of high active microbial blends, to achieve high efficiency and low consume, reduce equipment cost, enhance running stabilization of reactor. The adaptability test of microbial blends on different wastewater was carried on firstly. Oxygen consumption rate and anaerobic specific activity of methane producing of 15 different microbial blends were measured separately taking glucose artificial wastewater, Chinese herb extracting wastewater, brewery wastewater and ammonia nitrogen artificial wastewater as substrate, by which the adaptabilities of different microbial blends to wastewater were compared. According to the results high active microbial blends 14#, 8# and 10# were selected and used in the continuous treatment of wastewater in the laboratory and had obtained good effect, which had laid a foundation for application microbial blends to small scale test and pilot test of integrated anaerobic-aerobic bio-reactor. After the small scale test, the pilot test of the integrated anaerobic-aerobic bio-reactor treating fermentation wastewater was carried on. The test results showed fast initiation of the reactor. When system organic load reached 2.72 kgCODm-3d-1the COD removal rate of the reactor was stable between 84.5%～93.19% and it initiated successfully in more than 30 days at a time. In the load shock test the maximum organic load of system could reach to 8.88 kgCODm-3d-1 and the COD removal rate could be stable between 88.10%~96.88% which indicated that the reactor was efficient for treating wastewater and had strong resistance to shock load. At stable running period the COD removal rate of the reactor was over 90% and each index of wastewater could reach to the national discharge standards. In addition, the high active microbial blends and the macromolecule compound granule carrier, the matching system of the reactor was studied. It showed that the microbial blends adapted well to the reactor and abundant and high active microbes were formed in each functional field. The TS of granule sludge in anaerobic field was as high as 83.9 gL-1, the VS/TS was 56.9%~57.4%, the specific activity of methane producing was 280~350 mLCH4 gvss-1d-1. And the TS of immobilized biological granule was as high as 1.921 gL-1, the VS/TS was 94.02%~94.30%. Study on the carrier showed that the self-made macromolecular compound granule carrier was moderate of density, easy of fluidization, unease of running off, rough and porous, easy of films fixation, no bio-toxic, stable and safe, was a kind of superior carrier. Analysis of degradation process in each functional field confirmed the reactor, microbial blends and carriers were in good adaptation and wastewater was decontaminated by their cooperation.

过量Mg~(2+)对农作物种子萌发特性的影响(英文)

[Objective] The research aimed to reveal the effects of excessive Mg2+ on the germination characteristics of maize and soybean. [Method] Eleven concentration gradients of Mg2+ including 0,40,80,120,160,200,240,280,320,360 and 400 mmol/L were set up to study the effects of different treatments on such indices as the germination potential,germination rate,fresh weight,dry weight,root length,plant height and stem diameter of maize and soybean. [Result] With the increasing of Mg2+ concentration,the germination of maize delayed and the germination rate of soybean obviously decreased,reaching the significant difference at 0.05 level. When Mg2+ concentration increased,the fresh weight and dry weight of maize and soybean decreased,but maize and soybean showed some adaptability. The growth of radicles and seedlings in maize and soybean were obviously inhibited by Mg2+,and atrophy and dysplasia phenomena appeared. [Conclusion] Excessive Mg2+ stress has different effects on the germination potential and germination rate of different crops and has obvious poisoning effects on the root and stem growth of crops.

On-line voltammetric monitoring of dissolved Cu and Ni in the Gulf of Cadiz, south-west Spain

Geochemical processes in estuarine and coastal waters often occur on temporally and spatially small scales, resulting in variability of metal speciation and dissolved concentrations. Thus, surveys, which are aimed to improve our understanding of metal behaviour in such systems, benefit from high-resolution, interactive sampling campaigns. The present paper discusses a high-resolution approach to coastal monitoring, with the application of an automated voltammetric metal analyser for on-line measurements of dissolved trace metals in the Gulf of Cadiz, south-west Spain. This coastal sea receives metal-rich inputs from a metalliferous mining area, mainly via the Huelva estuary. On-line measurements of dissolved Cu, Zn, Ni and Co were carried out on-board ship during an eight-day sampling campaign in the study area in June 1997. A pumping system operated continuously underway and provided sampled water from a depth of ca. 4 m. Total dissolved metal concentrations measured on-line in the Gulf of Cadiz ranged between <5 nM Cu (<3 nM Ni) ca. 50 km off-shore and 60–90 nM Cu (5–13 nM Ni) in the vicinity of the Huelva estuary. The survey revealed steep gradients and strong tidal variability in the dissolved metal plume extending from the Huelva estuary into the Gulf of Cadiz. Further on-line measurements were carried out with the automatic metal monitor from the bank of the Odiel estuary over a full tidal cycle, at dissolved metal concentrations in the μM range. The application confirmed the suitability of the automated metal monitor for coastal sampling, and demonstrated its adaptability to a wide range of environmental conditions in the dynamic waters of estuaries and coastal seas. The near-real time acquisition of dissolved metal concentrations at high resolution enabled an interactive sampling campaign and therefore the close investigation of tidal variability in the development of the Huelva estuary metal plume.

干旱胁迫下油菜素内酯对文冠果苗木抗氧化酶活性和抗氧化剂含量的影响

用蘸根和叶面喷施不同浓度油菜素内酯(BRs)的方法,在人工控制土壤水分条件下,对黄土高原重要造林树种文冠果苗木重要的抗氧化酶活性和抗氧化剂含量变化进行研究。结果表明,用0.05~0.4 mg/L BRs处理文冠果苗木,超氧化物歧化酶(SOD)活性在中度干旱胁迫下较清水对照增加,但差异不显著,重度胁迫下增加显著;在中度和重度胁迫下,各处理文冠果苗木过氧化氢酶(CAT)、过氧化物酶(POD)和抗坏血酸过氧化物酶(APX)活性均较清水对照升高,且差异显著;在中度和重度胁迫下,5个浓度BRs处理对抗坏血酸(ASA)和还原型谷光甘肽(GSH)含量均具显著的增加效应。轻度和重度胁迫下,0.2 mg/L BRs处理对文冠果苗木抗氧化酶活性和抗氧化剂含量的增加效应均最为显著。

小麦抗旱生态分类中适合性聚类方法的研究

探索了适合于小麦品种抗旱生态分类的聚类方法 .选用 2 1个农艺性状和 15个冬小麦品种 (系 ) ,在聚类分析的各环节上 ,通过采用不同的策略 ,大规模进行了各种分类结果的比较 .结果表明 ,在与专家经验分类接近程度上 ,数据转换方法中 ,原始数据法依次大于普通相关阵基础上的方差极大正交旋转法、Promax斜交旋转法、主成份法 ;相似性度量上 ,欧氏距离大于马氏距离 ;聚类方式上 ,对应分析法和模糊聚类法大于最短距离法、最长距离法、类平均法 ;所有可组合的方法中 ,以对应分析法和直接用原始数据的模糊聚类法的分类结果最接近专家经验分类 .结合各方法理论上优缺点的分析与检验 ,认为这两种方法也是较理想的方法 .

干湿变化与作物补偿效应规律研究

干湿变化是作物生产的水分环境,利用植物旱后复水所产生的补偿效应是农业抗旱节水的新途径。论述了农业干湿变化类型和补偿效应的内涵、干湿变化对作物生态补偿和生长发育阶段间补偿及生理代谢功能间补偿性。通过土壤大气湿度组合的玉米实验,研究了大气湿度提高补偿土壤干旱作物生长与水分利用效应的规律。

黄土高原丘陵沟壑区沟道水资源利用模式初探

黄土高原丘陵沟壑区干旱缺水 ,要让有限的水资源支撑当地的生态农业建设 ,就要高效利用有限水资源 ,提高单位水的利用效率。通过在延安市燕儿沟的生产实践证明 ,在沟道中兴建截潜流工程 ,引水上山 ,发展坡地果园和保护地蔬菜微灌 ,是高效利用当地有限水资源的一种重要模式 ,很有推广价值

西部大开发中退耕还林(草)面临的挑战及其作用

根据中国西部森林的现状 ,采用生态学与地理学的方法 ,分析了因毁林开荒造成水土流失、洪涝灾害、沙尘风暴、干旱少雨、江河断流、物种减少等危害对国民经济发展所造成的影响 ,以及近几年所产生的一系列生态环境问题 .探讨了退耕还林 (草 )对我国的生态环境治理、西部持续发展、江河整治、国土整治综合措施运用、西部农民脱贫致富的作用 .

21世纪中国水资源危机

分析国内外水资源的现状、中国水资源危机及走向 2 1世纪存在的主要水问题 .说明中国属于世界上贫水的国家之一 ,人均拥有量只占世界平均水平的 2 5% ,在 6 6 8座城市中有 4 0 0余座缺水 .进入 2 1世纪 ,中国的水资源矛盾将进一步加剧 ,洪灾、干旱、断流、水污染、水土流失等问题将继续存在 .中国必须进行大规模国土整治 ,逆转恶化的水生态环境 ,合理开发、利用、保护水资源 .

不同倍性小麦材料对水分和密度条件的响应

以栽培一粒(2n)、栽培二粒(4n)和长武134(6n)为材料在大型活动防雨棚条件下研究不同倍性小麦材料在不同密度和水分条件下的产量适应性变化。结果发现,在两种水分条件下随着染色体倍性从2n→6n的增加,产量、千粒重、水分利用效率(WUE)和收获指数均呈增加趋势,在水分胁迫下各材料穗粒数和穗数则呈降低趋势,而在正常供水下穗粒数则呈增加趋势。在水分胁迫下栽培一粒、栽培二粒和长武134最高产量分别出现在中、低、高密度群体,而同一材料不同密度群体间变异系数分别为6.73%,1.98%,9.07%;不同倍性材料千粒重均随着密度增加而减小,而穗数则逐渐增加,二倍体的穗粒数以中密度最高,四倍体的穗粒数随着群体密度的增加而减小,六倍体则相反;三种材料WUE和收获指数分别以低、高、低密度最高。正常供水下随着染色体倍性从2n→6n的增加,三个倍性材料最高产量分别出现在高、低、低密度群体,而同一材料不同密度群体间变异系数分别为6.01%,17.12%,2.46%;千粒重表现为中密度>低密度>高密度,而穗粒数均以低密度群体最高,二倍体和四倍体以高密度群体最低,六倍体则以中密度群体最低,穗数则随着密度群体增加而增加;二倍体WUE以高...

辽西半干旱地区冬灌后覆盖的抗旱增产作用

辽西半干旱地区冬灌后覆盖的抗旱增产作用王仕新，庄季屏（中国科学院沈阳应用生态研究所，１１００１５）ＥｆｆｅｃｔｏｆＭｕｌｃｂｉｎｇＡｆｔｅｒＷｉｎｔｅｒＩｒｒｉｇａｔｉｏｎｏｎＤｒｏｕｇｂｔＲｅｓｉｓｔａｎｃｅａｎｄＹｉｅｌｄＩｎｃｒｅｍｅｎｔｉｎＳ...

浅论黄土高原集雨补灌农业的地位与作用

干旱缺水与水土流失并存是制约黄土高原经济发展的两大瓶颈性因素.集雨补灌农业作为雨水利用的更高发展阶段,更加强调了从时间和空间2个方面对有限雨水资源实施主动调控与利用.大量研究与实践证明,集雨补灌农业不仅是黄土高原不可缺少的水资源开发利用形式,也是黄土高原水土保持工程技术体系的重要组成部分,更是黄土高原旱地农业持续发展的一种综合模式和战略性措施,是对旱地农业的进一步继承和发展.另外,具有工程化、科技化、规模化内涵的集雨补灌农业也已经成为现代节水农业技术体系的重要研究内容之一.黄土高原集雨补灌农业研究的深度和广度将会持续深入,其技术发展更加依赖于高新技术的支撑与应用.

挖掘作物抗旱节水潜力——作物对多变低水环境的适应与调节

本文针对半干旱地区多变低水的实际田间环境,讨论了作物对不同干旱类型的反应,适度干旱后复水的生理补偿效应,不同类型作物在干湿交替条件下的差异,以及增强作物对多变水环境适应的技术.同时,就多变低水环境的概念、作物对多变低水环境的适应机理以及作物抗旱生理研究与旱农生产实际相衔接等问题提出了若干看法.

黄土丘陵沟壑区乔灌木植物系数计算与适应性评价

利用长期定位试验资料,研究了黄土高原丘陵沟壑区乔木和灌木植物系数的计算方法,比较了两种代表性乔木(榆树和小叶杨)和两种代表性灌木(柠条和沙棘)的实际蒸散量和生育期土壤平均含水量的差异,分析了土壤含水量变化对植物根系吸水和土壤水分有效性的影响。结果发现:该地区灌木的实际蒸散量小于乔木,但同类植物的实际蒸散量没有显著差异;土壤含水量柠条最高,沙棘最低;榆树植物系数最小,柠条其次,沙棘最大,但榆树和小叶杨的土壤水分有效性高,耗水量大,所以灌木较适于该区生长。