Implications of N acquisition from atmospheric NH3 for acid-bAse and cation-anion balance of Lolium perenne

In the atmosphere, ammonia (NH₃) is the third most abundant N species which, due to various natural and anthropogenic sources, can locally reach high concentrations. The acquisition of atmospheric NH₃ by plant shoots will lead to two opposing effects on acid-base balance. Absorption and dissolution of NH₃ will cause an alkalinisation, while the assimilation of NH₃ results in an acidification. Different rates of these processes would lead to an acid-base imbalance with consequences for the ionic balance of the plant. As there is only a limited capacity for biochemical disposal of excess H⁺ in shoots, pH regulation may involve a pattern of (in)organic ion flow between shoots and roots followed by H⁺/OH^? extrusion into the media via roots. The acquisition of NH₃ as additional N source should lead to a reduction in the ratio of mol H⁺/OH^? gained per mol N assimilated. We have recently investigated the NH₃ acquisition by Lolium perenne L. cv. Centurion and studied the effects of gas phase NH₃ on growth, acid-base balance and water-use efficiency. The experiments, therefore, included the application of a range of ¹⁴NH₃ to the shoots and of ¹⁵N as NO₃^?, NH₄⁺ or NH₄NO₃ to the roots. After a summary of the main conclusions from those experiments, we discuss the implications of the use of atmospheric NH₃ for the mineral composition of the plants. Over the range of NH₃ supplied, plants from all treatments could utilize gas-phase NH₃. Plants receiving NO₃^? via their roots had a higher capacity to use gaseous NH₃ than those growing with NH₄⁺. NH₃ assimilation in shoots reduced both the acid load with NH₄⁺ nutrition and the alkaline load with NO₃^? supply to the roots. The most significant effect of fumigation on the ion balance was an increase in K⁺ within all treatments, and this effect was highest in the NH₄⁺-fed plants. The results of the experiments support predictions of a combination of neutralizing biochemical reactions as well as transport of organic anion salts between shoots and roots as possible acid-base regulation mechanisms of the whole plant.     

Nitrogen assimilation in Lolium perenne colonized by the arbuscular mycorrhizal fungus Glomus fasciculatum

To investigate nitrogen assimilation in Lolium perenne L. colonized by the arbuscular mycorrhizal (AM) fungus Glomus fasciculatum (Thax. sensu Gerd.), nitrate uptake, key enzyme activities, and ¹⁵N incorporation into free amino acids were measured. After a 4-h labelling period with [¹⁵N]nitrate, ¹⁵N content was higher in roots and shoots of AM-plants than in those of control plants. Glutamine synthetase (GS) and nitrate reductase (NR) activities were increased in shoots of AM-plants, but not in roots. More label was incorporated into amino acids in shoots of AM plants. Glutamine, glutamate, alanine and γ-aminobutyric acid were the major sinks for ¹⁵N in roots and shoots of control and AM plants. Interactions between mycorrhizal colonization, phosphate and nitrate nutrition and NR activity were investigated in plants which received different amounts of phosphate or nitrate. In shoots of control plants, NR activity was not stimulated by high levels of phosphate nutrition but was stimulated by high levels of nitrate. At 4 m M nitrate in the nutrient solution, NR activity was similar in control and AM plants. We concluded that mycorrhizal effects on nitrate assimilation are not mediated via improved phosphate nutrition, but could be due to improved nitrogen uptake and translocation.     

农杆菌介导GUS基因对多年生黑麦草转化的研究

通过检测愈伤组织中GUS基因的瞬间表达,研究农杆菌LBA4404/pCAMBIA1301介导多年生黑麦草的转化体系。通过对多年生黑麦草瞬间表达率的比较,确立了其遗传转化的最佳优化条件。研究发现,多年生黑麦草不同品种的转化率在25%~45%之间变化。多年生黑麦草遗传转化最佳优化条件是预培养10d的胚性愈伤组织、浓度为0.5~0.8OD的农杆菌菌液以及2d共培养时间。在共培养基中添加100μmol/L乙酰丁香酮能有效地提高植物瞬间表达率。两种侵染处理方法比较结果为滤纸滴加法比浸泡法更优。转化后对愈伤组织的干燥处理能抑制农杆菌过度繁殖,能改善愈伤状态,有利于提高转化率。     

Detection of fungal infection in Lolium perenne by Fourier transform infrared spectroscopy

Aims The goal of the study was to apply Fourier transform infrared (FTIR) spectroscopy followed by chemometrical data treatment for the differentiation of fungi-infected perennial ryegrass (Lolium perenne) from uninfected grass.Methods FTIR was used to rapidly discriminate between leaves of perennial ryegrass (L. perenne) infected by a fungal endophyte (Epichlo?; asexual forms: Neotyphodium) and uninfected leaves. Besides drying and grinding of the sampled leaves, no other preparation steps were needed. FTIR measurements were performed in the attenuated total reflection (ATR) mode. Aliquots of powdered leaf samples were placed on a ZnSe crystal and the spectra were collected, followed by chemometrical analysis (multidimensional factor analysis, hierarchical cluster analysis).Important findings ATR-FTIR allowed a rapid detection of fungal infections in the plant material and proved to be a fast and reliable tool for the differentiation of plant biomass without the need of time-consuming sample preparation.     

多年生黑麦草种子萌发及植株再生的研究

对不同预处理条件下多年生黑麦草种子萌发、愈伤组织诱导以及植株再生进行了研究。结果表明,35℃恒温干燥处理3 h、浸泡16 h后,室温晾48 h的干湿交替及强度为1 000 GS磁场处理30 min均能使种子的发芽率得到显著提高。最适诱导培养基为MS+2,4-D 6 mg·L^-1,有机添加物对于愈伤组织的生长影响并不显著,但可以改善其状态;较为适宜的分化培养基为MS+2 mg·L^-1 6-BA或KT;生根培养基为附加0.5 mg·L^-1 NAA的MS培养基。     

多年生黑麦草抗氧化酶和植物络合素对Cd2+胁迫的应答

采用水培方法研究了5 mg· L-1 Cd2+胁迫下,Cd在多年生黑麦草中的积累和Cd2+对多年生黑麦草抗氧化酶活性和植物络合素等巯基化合物浓度的影响.将具有3片展开叶的多年生黑麦草实生苗转至1/2霍格兰营养液中培养2周后,对其进行5 mg-L-1 Cd2+处理,分别在处理后的0、0.25、1、3、6d取样测定根系和叶片的Cd浓度、抗氧化酶活性和植物络合素等巯基化合物的浓度.结果表明,Cd2+处理多年生黑麦草6d后,根系中Cd浓度达到2.59 mg·g-1,叶片中Cd浓度达到0.24 mg·g-1,根中Cd向叶片的转运系数力0.093,叶中Cd的富集系数为48,多年生黑麦草属Cd高积累植物,具备在植物修复上应用的前景.Cd2+胁迫下,多年生黑麦草根叶中丙二醛(MDA)含量无显著变化,根中超氧化物歧化酶(SOD)、过氧化氢酶(CAT)活性无显著变化,抗坏血酸过氧化物酶(APX)对Cd2+敏感,处理后6d活性较0d显著下降67.19％.Cd2+处理1d内,叶中SOD、APX、CAT活性显著降低.Cd2+处理后3d,叶中的抗氧化酶系统对叶中Cd浓度的升高做出了正反馈,SOD、APX、CAT的活性分别较处理后1d显著上升了14.19％、76,82％、99.26％,Cd2+处理时间延长至6d,SOD活性较处理后3d显著下降了18.58％,APX、CAT活性无显著变化.Cd2+处理后6d,多年生黑麦草根中半胱氨酸(Cys)、谷胱甘肽(GSH)、植物络合素2(PC2)、植物络合素3(PC3)、植物络合素4(PC4)、植物络合素5(PC5)和植物络合素6(PC6)浓度分别较处理0d提高了2.19、1.57、2.06、16.08、5.73、6.03和4.31倍,叶中Cys、GSH、PC2、PC3和PC4浓度分别较处理0d提高了0.69、3.21、1.64、5.73和0.27倍.根中PC3巯基比例最大,叶中GSH的巯基比例最大,二者是根、叶中巯基存在的主要形式.随着Cd2+处理时间的延长,根系和叶片中各巯基化合物的总巯基浓度显著升高,根系和叶片中植物络合素总巯基浓度与Cd浓度显著正相关.多年生黑麦草通过植物络合素等巯基化合物的快速合成降低了根叶中自由Cd2+的比例,保护了根叶中抗氧化酶的活性,间接维持了活性氧代谢的平衡.     

UV-B and PAR in a grass (Lolium perenne L.) canopy

The penetration of natural and artificial UV-B_BE (Biologically Effective UV-B, Caldwell 1971) and PAR (400–700 nm) in a grass canopy with increasing LAI was followed during 2 months. Overall, the transmission of UV-B_BE sunlight is significantly higher than of PAR sunlight. This is mainly due to the higher proportion of diffuse light in the UV-B. Under cloudy conditions no difference between UV-B_BE and PAR could be found. Sun angle and intensity of the radiation were less important in determining the penetration of light. Artificial light penetrates much more through the canopy, resulting in higher irradiation levels in the lower part of the canopy, but a lower UV-B_BE/PAR ratio (since UV-B transmittance of the leaves is lower). The UV-B_BE/PAR ratio reaching the leaves was influenced by LAI, sun angle, percent diffuse light and leaf angle. The large differences in UV-B_BE/PAR ratio per unit leaf area under natural and artificial light conditions are important in understanding the influence of UV-B on plants.     

Absorption of nitrate and ammonium ions by Lolium perenne from flowing solution cultures at low root temperatures

Lolium perenne L. cv. 23 (perennial ryegrass) plants were grown in flowing solution culture and acclimatized over 49 d to low root temperature (5°C) prior to treatment at root temperatures of 3, 5, 7 and 9°C for 41 d with common air temperature of 20/15°C day/night and solution pH 5·0. The effects of root temperature on growth, uptake and assimilation of N were compared with N supplied as either NH₄ or NO3 at 10 mmol m^?3. At any given temperature, the relative growth rate (RGR) of roots exceeded that of shoots, thus the root fraction (Rf) increased with time. These effects were found in plants grown with the two N sources. Plants grown at 3 and 5°C had very high dry matter contents as reflected by the fresh weight: freeze-dried weight ratio. This ratio increased sharply, especially in roots at 7 and 9°C. Expressed on a fresh weight basis, there was no major effect of root temperature on the [N] of plants receiving NHJ but at any given temperature, the [N] in plants grown with NHJ was significantly greater than in those grown with NO₃. The specific absorption rate (SAR) of NH⁺₄ was greater at all temperatures than SAR-NO₃. In plants grown with NH⁺, 3–5% of the total N was recovered as NH⁺₄, whereas in those grown with NO^?₃ the unassimilated NO^?₃ rose sharply between 7 and 9°C to become 14 and 28% of the total N in shoots and roots, respectively. The greater assimilation of NH⁺₄ lead to concentrations of insoluble reduced N (= protein) which were 125 and 20% greater, in roots and shoots, respectively, than in NO^?₃-grown plants. Plants grown with NH⁺₄ had very much greater glutamine and asparagine concentrations in both roots and shoots, although other amino acids were more similar in Concentration to those in NO^?₃ grown plants. It is concluded that slow growth at low root temperature is not caused by restriction of the absorption or assimilation of either NH⁺₄ or NO^?₃. The additional residual N (protein) in NH⁺₄ grown plants may serve as a labile store of N which could support growth when external N supply becomes deficient.     

黑麦草内生真菌感染状况的检测及定量分析

选取内生真菌特异性引物,成功建立了利用PCR技术对黑麦草中内生真菌感染状况的检测和定量分析方法。此检测方法的准确性高于常规乳酸-苯胺蓝染色法。利用实时荧光PCR定量分析的结果表明：不同植株之间内生真菌含量差异较大,而同株植物相同龄级分蘖之间内生真菌含量无显著差异。由此可见内生真菌的含量不仅与植物种以及品种有关,也与植物的基因型密切相关。     

重金属递进胁迫对黑麦草初期生长的影响

通过研究Cu²⁺、Zn²⁺、Cd²⁺与Pb²⁺胁迫对黑麦草初期生长的影响,结果表明：4种重金属对种子发芽率抑制效应相对较小,尤其Cu²⁺与Zn²⁺的抑制作用最小。高浓度Cu²⁺、Cd²⁺胁迫对株高、根系长度、地上生物量的抑制作用相对较大,尤其Cu²⁺对根系生长的抑制效应最大,在300 mg·L^-1下,与对照相比,根长最高下降了 85.48%。高浓度Cd²⁺胁迫显著降低了叶绿素含量,在300 mg·L^-1时比对照降低了45.51%;与对照相比,Cu²⁺与Zn²⁺所有处理都增加了叶绿素含量。从递进胁迫进程看,一些重金属对某一生长指标的影响往往表现在低浓度具有促进作用,而高浓度又存在明显的抑制效应。     

Modelling the spread of pollen from Lolium perenne. The implications for the release of wind-pollinated transgenics

The dispersal of pollen from a Lolium perenne source has previously been described using various Gaussian plume models which take distance and wind direction into account. One of these models is used here to calculate, using integration, possible pollen deposition onto small conspecific populations a kilometer from the source. The percentage of immigrant pollen is compared for six different sets of parameter values previously estimated from pollen-dispersal experiments. The source size is then scaled up to simulate what might happen if transgenic ryegrass was grown on a large scale. In this case it is seen that small conspecific populations might, in some conditions, be swamped by immigrant pollen, even if they are not directly downwind of the source. The implications of this are discussed in terms of assessing and managing the risks of releasing wind-pollinated transgenic crops. Received: 10 July 1999 / Accepted: 27 August 1999     

紫茎泽兰与黑麦草苗期相对竞争力和生长特征的研究

为探究替代物种黑麦草（Lolium perenne）抑制外来人侵杂草紫茎泽兰（Eupatorium adenophorum）的潜力,采用de Wit取代系列设计,在野外小区内设置3水平多年生黑麦草栽植时间、4水平种植密度以及3水平混种比例等不同种植处理,于12周后收获,分别测量黑麦草与紫茎泽兰的地上生物量和地下生物量,用单株相对产量分析其竞争力。结果表明,种植密度、混种比例和黑麦草栽植时间显著影响紫茎泽兰的地上生物量,紫茎泽兰的竞争力在低种植比例情况下,随黑麦草栽植时间的推迟而明显增强,随种植密度的降低而增强。种植密度、混种比例、黑麦草栽植时间以及这些因素的互作等对黑麦草的地上生物量与地下生物量均具有显著影响;栽植时间越晚、密度越大,黑麦草的竞争力就越弱（尽管不存在直线关系）。在大多数种植处理条件下,紫茎泽兰经历的种间竞争明显大于种内竞争,表现出较弱的竞争力;而黑麦草在与紫茎泽兰同时栽植的情况下,经历的种内竞争大于种间竞争,表现出较强的竞争力。     

土壤干旱条件下保水剂对多年生黑麦草光合特性的影响

以多年生黑麦草（Lolium perenne Linn．）为研究对象,对土壤干旱条件下（土壤含水量12．6％、10．5％、8．4％和6．3％）使用保水剂后叶片光合特性的动态变化进行了分析。结果显示：随土壤含水量的降低和胁迫时间（12d）的延长,多年生黑麦草叶片的净光合速率（Pn）、气孔导度（Gs）、蒸腾速率（Tr）和光能利用效率（LUE）呈逐渐下降的趋势;随土壤含水量的降低,胞间CO2浓度（Ci）逐渐升高、瞬时水分利用效率（WUE）逐渐降低,但随胁迫时间的延长Ci和WUE则呈现不同的变化趋势;总体上,在土壤含水量不同的条件下及不同的胁迫时间各指标均有显著差异（P〈0．05）。与未添加保水剂的各处理组相比,添加质量分数2％保水剂后多年生黑麦草叶片的Pn、Gs、WUE和LUE值均增大,而Ci和Tr值则降低,差异达显著水平（P〈0．05）。研究结果表明：合理使用保水剂能提高多年生黑麦草叶片的光合能力以及土壤的保水能力,增强多年生黑麦草对干旱环境的适应性。     

不同氮素营养条件下内生真菌感染对黑麦草光合和蒸腾速率及生物量的影响

以黑麦草(Lolium perenne L)为实验材料,研究在不同氮素营养条件下内生真菌(Neotyphodium lolii)感染对其光合特性及生物量的影响.结果表明:1)整个实验中,内生真菌感染对黑麦草净光合速率、蒸腾速率以及水分利用效率均无显著影响,但在中氮和高氮水平时,感染种群的净同化累积值有超过非感染种群的趋势;2)在本实验的第一处理期和第二处理期,内生真菌感染对黑麦草绿色部分的干物质产量无显著影响,但在第三处理期感染种群的绿色部分的干物质产量显著高于非感染种群.     

黑麦草(Lolium perenne L.)内生真菌的检测、分离及鉴定

从多年生黑麦草(LoliumperenneL.)5个品种———SR4000、Pinnacle、Topgun、CalypsoⅡ、Justus中分离出61个菌株。次培养后,所得形态稳定的菌株可分为4个形态群,依据其形态特征及APPCR的结果,确定其中的57个分离菌株为Neotyphodiumlolii。