Evidence For Root Contraction In White Clover (Trifolium repens L.)

White clover (Trifolium repens L.) stolons become buried inthe field. It was observed that this also occurred in the greenhousewhere the accepted mechanisms of burial, treading by livestockand earthworm casting, did not occur. It was also observed thatthe crown of seedling T. repens plants become closely appressedto the soil. Experiments showed that, regardless of varietyof T. repens or depth of planting, all seedling hypocotyls firstlift the cotyledons clear of the soil, then ‘ contract’towards the soil until the cotyledons are in contact with orbelow the soil surface. Auxanometers were used to measure therate and extent of this contraction and were also attached tostolon nodes in experiments which showed that stolons move downwardsrelative to the soil surface and that the speed and extent ofthis duration varied with soil type. A further experiment showedthat only rooted nodes show this behaviour. The force exertedby the contraction of nodal roots was estimated experimentallyas 0.21 N g^-1fresh root. A mechanism for the root contraction,based on examination of root anatomy of seedling tap-roots andnodal roots, is suggested. These experiments provide evidencefor root contraction in T. repens which may lead to stolon burial.The importance of this to T. repens as a pasture species andas a means of further improving T. repens varieties is discussed.Copyright 1999 Annals of Botany Company White clover, Trifolium repens, L., stolon, seedling, burial, root, nodes, nodal, force, contractile, soil resistance, pasture, phloem, fibres.     

利用P5CS基因转化白三叶的研究

为了培育抗旱白三叶,构建了植物表达载体pBPC-P5CS,利用基因枪法转化白三叶的愈伤组织,PCR检测和Southern blot鉴定证实白三叶中已导入P5CS基因。对转P5CS基因白三叶植株的不同抗旱指标进行了分析。结果表明,与对照相比,转P5CS基因株系的抗旱能力得到了较大的提高。干旱胁迫下,与对照相比,转P5CS基因植株的脯氨酸含量和相对含水量分别比对照高20.0%-21.2%和5.6%-8.5%。     

Floral Initiation of White Clover (Trifolium repens L.) in Response to Daylength and Temperature

The response to different combinations of daylength (14, 15and 16 h) and day/night temperature (20/10°C, 30/15°C)of four white clover (Trifolium repens) varieties in terms offloral initiation and development was studied. More plants flowered under long days (16 h) at 30°C thanin any other combination. Varietal differences in response todaylength were large but temperature was the factor controllinginitiation. Daylength was more important in regulating peduncle length,floret and ovule numbers than was temperature. The amount ofnectar produced was lowest at the higher temperature althoughnectar concentration was unaffected by temperature. Nectar concentrationwas lowest in the shortest daylength (14 h). White clover, floral initiation, temperature, daylength     

Phosphate Uptake and Transport by Roots and Stolons of Intact White Clover (Trifolium repens L.)

The extent to which phosphate can be absorbed directly fromthe outer medium by stolon internodes and contribute to thetotal accumulation of phosphate by intact plants of white clover(Trifolium repens L. cv. Blanca) was assessed in hydroponicexperiments in a controlled environment room. The uptake ofphosphate by intact roots or stolons was measured by sealinga segment (6-0 mm long) across a flow-cell in which ³²P-labellednutrient solution was circulated for 24 h, the rest of the rootsystem receiving unlabelled nutrient solution. The rate of uptakeof phosphate (µmol g^–1 d^–1 dry wt. basis)by roots was more than 300 times that by intact stolons. Pretreatmentof stolons by gentle abrasion to remove cuticle, so as to simulatethe condition of stolons in the field, increased the uptakeof phosphate 7-fold compared with that of intact stolons. However,the potential of stolons to contribute to the P status of whitedover in the field was calculated to be small (5%). When an incision was made through the hypodermal layer of stolons,the rate of phosphate uptake greatly increased, attaining 71%of that by root segments. This increase, which was greater athigher phosphate concentrations, indicates that the suberi.zedhypodermis constitutes a major barrier to the influx of phosphatein the stolon. After withholding phosphate for different time intervals, thesubsequent rate of phosphate uptake by roots was increased 2-3-foldafter 2 d phosphate deprivation and 3-4-fold after 6 d or 13d phosphate deprivation. A higher proportion of absorbed phosphatewas transported to shoots in phosphate-deprived plants. After1 d of uptake following restoration of the phosphate supply,the concentrations of labelled phosphate in shoots were greaterthan in control plants, although the concentrations of labelin roots was less. However, the rate of uptake of phosphateby stolons, following deprivation, was not significantly increased.These results suggest that the mechanism regulating the enhancedrate of phosphate loading into the xylem, initiated by a periodof phosphate deprivation, is specific to roots and is not inducedin stolons. The results are discussed in relation to the growth and acquisitionof phosphate by white clover in the field. Key words: Nutrient deficiency, phosphate, stolons, transport (ions), Trifolium repens     

3D Architectural Modelling of Aerial Photomorphogenesis in White Clover (Trifolium repens L.) using L-systems

The objective of this work was to construct a model of aerialdevelopment of clover that takes into account morphogeneticresponses to the light environment, and to use it to analyseand understand these processes in terms of signal perceptionand integration. The plant model was interfaced with a MonteCarlo model that determines photosynthetically active radiation(PAR) and red/far-red ratio (R/FR) throughout the canopy, takinginto account the absorption, reflection and transmission oflight by individual leaves. Light intensity and quality weresensed by the plant model at discrete time intervals and atdiscrete sites of perception: apices, emerging internodes andpetiole tips. This input regulated the final size of internodesand leaves, the vertical positioning of leaves, and the branchingdelay. The empirical relations (regression functions) quantifyingthis regulation were derived from data reported in the literatureand original measurements. Simulations produced realistic visualizationsand quantitative characterizations of the modelled plants fordifferent light treatments. These results were in general agreementwith observations of real plants growing under similar conditions,suggesting that the dependence of organ size and position onlight treatments can be regarded as an integration of the responsesof individual plant organs to their local light environment.The model is used to describe the regulation of branch appearanceand the impact of self-shading on plant morphogenesis as a functionof local light environment. Copyright 2000 Annals of BotanyCompany Clover, Trifolium repens L, photomorphogenesis, plant architecture, L-system, modelling, Monte-Carlo method, competition for light, red : far-red ratio, irradiance, light quality, leaf size, self-shading     

Scavenger Enzyme Activities in Subcellular Fractions of White Clover (Trifolium repens L.) under PEG-induced Water Stress

Scavenger enzyme activities in subcellular fractions under polyethylene glycol (PEG)-induced water stress in white clover (Trifolium repens L.) were studied. Water stress decreased ascorbic acid (AA) content and catalase (CAT) activity and increased the contents of hydrogen peroxide (H₂O₂), thiobarbituric acid reactive substances (TBARS) (measure of lipid peroxidation), and activities of superoxide dismutase (SOD), its various isozymes, ascorbate peroxidase (APOX), and glutathione reductase (GR) in cellular cytosol, chloroplasts, mitochondria, and peroxisomes of Trifolium repens leaves. In both the PEG-treated plants and the control, chloroplastic fractions showed the highest total SOD, APOX, and GR activities, followed by mitochondrial fractions in the case of total SOD and GR activities, whereas cytosolic fractions had the second greatest APOX activity. However, CAT activity was the highest in peroxisomes, followed by the cytosol, mitochondria, and chloroplasts in decreasing order. Although Mn-SOD activity was highest in mitochondrial fractions, residual activity was also observed in cytosolic fractions. Cu/Zn-SOD and Fe-SOD were observed in all subcellular fractions; however, the activities were the highest in chloroplastic fractions for both isoforms. Total Cu/Zn-SOD activity, the sum of activities observed in all fractions, was higher than other SOD isoforms. These results suggest that cytosolic and chloroplastic APOX, chloroplastic and mitochondrial GR, mitochondrial Mn-SOD, cytosolic and chloroplastic Cu/Zn-SOD, and chloroplastic Fe-SOD are the major scavenger enzymes, whereas cellular CAT may play a minor role in scavenging of O₂⁻ and H₂O₂ produced under PEG-induced water stress in Trifolium repens.     

Assessing the Geometric Structure of a White Clover (Trifolium repens L.) Canopy using3-D Digitising

A method was developed for assessing the three dimensional (3-D)geometric structure of white clover canopies. 3-D co-ordinatesof pre-defined points on leaves, petioles and stolons were measuredusing a Polhemus Fastrak electromagnetic 3-D digitiser. Digitisingprogressed downwards from the top of the canopy and plant partswere removed after they have been digitised. Leaflets were treatedas four quarter-ellipses, and petiole and stolons were treatedas cylinders. Leaf dimensions and areas calculated from 3-Dco-ordinates were within about 5% and 20% of direct measurementsmade with a ruler and a planimeter, respectively. Special softwareand freeware POV-Ray were used to reconstruct a virtual canopyfrom digitiser records and to calculate canopy characteristicssuch as leaf area index (LAI), petiole intersection area, andprofiles of leaflet areas and inclinations with height. It tookbetween 3 and 7 h to digitise 10 x 10 cm stands of clover andthe resulting information was considerably more comprehensiveand accurate than could have been obtained by the alternative‘point quadrat’ or ‘stratified clipping’methods.Copyright 2000 Annals of Botany Company White clover, Trifolium repens, geometric structure, leaf area, leaf angle, 3-D digitising     

The Growth and Carbon Allocation Patterns of White Clover (Trifolium repens L.) Plants of Contrasting Branching Structure

The growth, morphology and carbon allocation patterns of F1progeny white clover (Trifolium repens L.) plants selected foreither low (‘LBF’) or high (‘HBF’) frequencyof stolon branching were compared in two controlled-environmentexperiments. Selections from within both a small-leaved (‘GrasslandsTahora’) and a large-leaved (‘Grasslands Kopu’)clover cultivar were compared, and plants were grown under arelatively lenient defoliation treatment (expt 1) or under threelevels of defoliation seventy (expt 2). Carbon allocation patternswere measured by ¹⁴CO₂ pulse-chase labelling using fully unfoldedleaves on the main (parent) stolon. LBF and HBF displayed consistent differences in the selectedcharacter though, within cultivars, the difference between selectionswas most pronounced for Kopu. The selections developed fundamentallydifferent branching structures resulting from differences inbranching frequency, with total branch weight per plant averaging122 mg for LBF and 399 mg for HBF (mean of both experiments).More C moved from parent stolon leaves to branches in HBF thanin LBF (mean 22.6% vs. 15.1% respectively of the ¹⁴C exportedfrom source leaves). More C also moved to stolon tissue in HBF,but, counterbalancing this and the difference in allocationto branches, less moved to developing leaves and roots on theparent stolon itself compared to LBF. However, the total weightof developing leaves and roots per parent stolon was generallygreater in HBF than in LBF, probably reflecting greater C importby these sinks from the higher number of branches present perplant in the former selection. HBF plants were consistentlylarger at harvest than LBF plants. There were no defoliationtreatment x selection interactions in C allocation patternsin expt 2. The implications of the results for plant performancein grazed pastures are discussed. Branching, carbon translocation, defoliation, growth, morphology, Trifolium repens, white clover     

Current Nitrogen Fixation Is Involved in the Regulation of Nitrogenase Activity in White Clover (Trifolium repens L.)

Previous studies have shown that nitrogenase activity decreases dramatically after defoliation, presumably because of an increase in the O2 diffusion resistance in the infected nodules. It is not known how this O2 diffusion resistance is regulated. The aim of this study was to test the hypothesis that current N2 fixation (ongoing flux of N2 through nitrogenase) is involved in the regulation of nitrogenase activity in white clover (Trifolium repens L. cv Ladino) nodules. We compared the nitrogenase activity of plants that were prevented from fixing N2 (by continuous exposure of their nodulated root system to an Ar:O2 [80:20] atmosphere) with that of plants allowed to fix N2 (those exposed to N2:O2, 80:20). Nitrogenase activity was determined as the amount of H2 evolved under Ar:O2. An open flow system was used. In experiment I, 6 h after complete defoliation and the continuous prevention of N2 fixation, nitrogenase activity was higher by a factor of 2 compared with that in plants allowed to fix N2 after leaf removal. This higher nitrogenase activity was associated with a lower O2 limitation (measured as the partial pressure of O2 required for highest nitrogenase activity). In experiment II, the nitrogenase activity of plants prevented from fixing N2 for 2 h before leaf removal showed no response to defoliation. The extent to which nitrogenase activity responded to defoliation was different in plants allowed to fix N2 and those that were prevented from doing so in both experiments. This leads to the conclusion that current N2 fixation is directly involved in the regulation of nitrogenase activity. It is suggested that an N feedback mechanism triggers such a response as a result of the loss of the plant's N sink strength after defoliation. This concept offers an alternative to other hypotheses (e.g. interruption of current photosynthesis, carbohydrate deprivation) that have been proposed to explain the immediate decrease in nitrogenase activity after defoliation.     

Developmental and Physiological Studies on the Cyanogenic Glucosides of White Clover, Trifolium repens L.

Cyanogenesis in Trifolium repens L. is under the control oftwo loci; Ac/ac and Li/Hi control cyanogenic glucoside and linamaraseproduction respectively. Results obtained show that neitherthe dominant allele (Ac) coding for cyanogenic glucoside productionnor the dominant allele (Li) coding for linamarase productionare expressed in roots, seeds or seedlings before shoot emergence.Both linamarase and cyanogenic glucoside are produced duringshoot growth and there is little turnover of cyanogenic glucosidein mature leaves. As the leaves senesce there is breakdown ofthe mechanism separating cyanogenic glucoside and linamarase,since cyanogenic glucoside is lost in plants of genotype AcAc Li Li but not in those of genotype Ac Ac Li Li. About 60%of the cyanogenic glucoside produced was lotaustralin, in shootsof plants which were fed with equal quantities of the precursoramino acids L-valine and L-isoleucine. In contrast, the proportionof cyanogenic glucoside as lotaustralin found in leaves of oneplant, was only 40%. Different plants were shown to producedifferent quantities of cyanogenic glucoside, and the amountproduced was dependent on temperature.     

Influence of Enhanced Concentration of Carbon Dioxide and Moderate Drought on Fluorescence Induction in White Clover (Trifolium repens L.)

In a 5-months' experiment with white clover grown in two CO₂ atmospheres (AC = 350 and EC = 700 cm³ m^-3) and two humidities (-0.2 and -0.8 MPa), changes in fluorescence induction parameters were investigated. Changes induced by EC did not coincide with those induced by moderate drought. Long-term EC significantly increased stationary fluorescence F_s and decreased the vitality coefficient Rfd. Moderate drought significantly shortened the fluorescence half-time and decreased the area above the induction curve S_c and the F_v/F_m ratio.     

Influence of Temperature and O2 Concentration on Photosynthesis and Light Activation of Ribulosebisphosphate Carboxylase Oxygenase in Intact Leaves of White Clover (Trifolium repens L.)

Detached leaves of white clover (Trifolium repens L.) were keptfor 1 h under various conditions of temperature, oxygen concentrationand light intensity. Rates of photosynthesis were measured whereappropriate and then ribulosebisphosphate carboxylase oxygenase(RuBPCO) was extracted rapidly and its initial activity measuredimmediately. The extracted activity increased with increased intensity ofillumination of the leaves. Where leaves were pretreated atlow light intensity, the lower the temperature of the leavesthe higher the extracted activity of RuBPCO. At high light intensitytemperature did not affect the activity of subsequently extractedRuBPCO but the light intensity which was necessary for maximumactivity increased with temperature. Activity of RuBPCO fromleaves pretreated in the dark was least when CO₂ was low andtemperature high. Leaves, pretreated at low temperatures andhigh light intensity in 20% O₂, yielded higher activity in extractsthan leaves pretreated under similar conditions but in 2% O₂.A relatively weak temperature response of photosynthesis atlow irradiances was associated with a decrease in extractableRuBPCO activity with increasing temperature. A strong temperaturedependence of the oxygen inhibition of photosynthesis was associatedwith lower extractable RuBPCO activity in leaves pretreatedat low oxygen concentration at low temperatures. With leavesfrom plants grown at low temperatures prior to treatment ofleaves, oxygen inhibition of photosynthesis was less temperaturedependent and activity of RuBPCO in extracts was not decreasedby low O₂ at low temperatures. Differences in the activationof RuBPCO appear to influence photosynthesis and account foran absence of oxygen inhibition of photosynthesis at low temperaturesin plants grown in warm conditions. Key words: Ribulosebisphosphate carboxylase oxygenase activation, Photosynthesis, Temperature, O₂ effect, White clover     

The Extent and Pattern of Osmotic Adjustment in White Clover (Trifolium repens L.) During the Development of Water Stress

White clover plants were subjected to water stress followingthe cessation of watering. As a water deficit developed, waterand osmotic potentials were measured in stolon tips, leavesfrom the stolon tip and leaves from the plant crown. Pressurepotentials were calculated. Pressure potential was maintainedin stolon tips even when water potential fell to around –2·0MPa. In contrast, pressure potential in leaves fell rapidlyas water stress developed. Total amino acid and potassium levels were largely unaffectedin both stolon tips and leaves. Water-soluble carbohydratesand proline accumulated during water stress. The increase inproline level in leaves did not follow the same pattern as thatin stolon tips, although toward the end of the water stressperiod the level had increased by a similar extent in both partsof the plant. Additionally, pressure potential and osmotic potentialappeared to be significantly related to proline content in stolontips. No such relationship was found for leaves. The role ofproline in osmotic adjustment is discussed. Trifolium repens L. cv. Olwen, white clover, water stress, osmotic adjustment, proline     

The Effect of Water Stress on Floral Characters, Pollination and Seed Set in White Clover (Trifolium repens L.)

White clover plants were subjected to either short-term developingwater stress or long-term stable levels of water deficit. Thehort-term stress reduced plant water status to about –2·0MPa over 16 d. The long-term stress was less severe, but wassustained for several weeks. Long-term water stress promotedthe production of inflorescences. However, water stress alsoincreased floret abortion and the premature death of whole flowerheads. The number of ovules per floret was decreased by waterstress. The most striking effect of both long- and short-term waterdeficit was to reduce pollen viability measured with the fluorochromaticassay. This was not an artefact of assay conditions. The pollenfrom water-stressed flower heads was not reversibly dehydrated;it did not score at similar viability to controls after incubationin conditions which hydrate pollen. In addition, the pollenfrom water-stressed plants lost viability more rapidly thanpollen from well-watered plants after removal from the flowerhead. The consequences of reduced pollen viability on seed set wereinvestigated by hand-crossing within and between groups of plantsmaintained for several weeks at three levels of water supply.Flower heads pollinated with pollen from water-stressed plantsset fewer seeds per floret than those pollinated with controlpollen. Key words: Trifolium repens, white clover, water stress, floral characters, seed set     

Influence of Shade on the Growth and Sink Activity of Young Flower Heads and Peduncles of White Clover (Trifolium repens L.)

The influence of shade on the growth and sink activity of youngflower heads and peduncles of white clover (Trifolium repensL.) plants was measured in controlled environments. Measurementof the peduncle elongation rate using a linear voltage displacementtransducer showed that when the inflorescence alone was shaded,peduncle elongation was higher than in the light. Translocationstudies using ¹¹C-labelling techniques showed that shading theinflorescence alone had little effect on translocation of assimilatesinto the inflorescence but induced a major change in partitioningof assimilates within it. Peduncle elongation induced by shadeaccompanied an increase in partitioning of photoassimilatesto the peduncle, at the expense of the flower head. Key words: White clover, partitioning, translocation     

Photosynthesis of White Clover (Trifolium Repens L.) Germplasms with Contrasting Leaf Size

In a growth chamber experiment, we determined net photosynthetic rate (PN) and leaf developmental characteristics of cultivars of a relatively small-, intermediate-, and a large-leaf genotype grown under irradiance of 450-500 µmol(photon) m-2 s-1 (HI), shade [140-160 µmol(photon) m-2 s-1] (LI), and after a shade-to-irradiation (LI »HI) transfer. Differences in physiological responses of the genotypes were more pronounced in HI and LI»HI plants than in LI plants. The small- and intermediate-leaf sizes had greater PN in the first measured leaf than the large-leaf type by 70 and 63 % in HI plants, and by 23 and 18 % in LI»HI plants, respectively. Similar relationships were observed in the next developed leaf. The LI plants did not differ significantly in PN. Greater PN in the small- and intermediate-leaf size genotypes were not associated with greater total dry matter of the plant. Under irradiation, the large-leaf genotype accumulated more total nonstructural saccharides (TNS) and starch than the small- or intermediate-leaf size plants. TNS and starch concentrations in LI plants were about one-half those of HI and LI»HI plants. These results should help to develop management practices that capitalize upon the competitive features of white clover in mixed-species swards.     

Effect of Carbohydrate Demand on the Remobilization of Starch in Stolons and Roots of White Clover (Trifolium repens L.) after Defoliation

White clover plants were grown from stolon tips in growth cabinetsand then defoliated. Thereafter, changes in the contents ofnon-structural carbohydrates such as starch, sucrose, glucose,fructose, maltose, and pinitol in stolons and roots were monitored.Initial contents of carbohydrate reserves, photosynthetic supplyof new carbohydrates and carbohydrate demand after defoliationwere varied by growing the plants at various CO₂ partial pressures,by varying the extent of defoliation and by removing eitherroots or stolon tips at the time of defoliation. Remobilization of carbohydrate reserves in stolons increasedproportionally to their initial contents and was greater whenplants had been severely defoliated, suggesting that carbohydrateswere remobilized according to availability and demand. Starchwas the predominant reserve carbohydrate. Starch degradationwas associated with decreased contents of sucrose, glucose andfructose in young stolon parts and roots but not in old stolonparts suggesting that starch degradation was not strictly controlledby the contents of these sugars. A decrease in the demand forcarbohydrates by removal of roots did not decrease starch degradationbut increased the contents of sucrose, glucose, and fructose.Removal of stolon tips decreased starch degradation and contentsof sucrose, glucose, and fructose. The results suggest thatstarch degradation was controlled by a factor other than sucrose,glucose, and fructose which was exported from stolon tips, e.g.gibberellin. Key words: White clover, storage carbohydrates, remobilization, regrowth     

Water Relations of White Clover (Trifolium repens): Water Potential Gradients and Plant Morphology

Water potential, osmotic potential, pressure potential and relativewater content were measured in stolons and leaves of white cloverplants grown under a range of conditions of water supply andevaporative demand. The importance of adventitious roots fromthe nodes was examined. Gradients along stolons were alwaysextremely small, of the order of only 01 MPa. Stolon up waterpotential was representative of plant water status regardlessof stolon length, presence/absence of nodal roots, degree ofwater stress and evaporative demand. It is concluded that waterconduction along stolons was very good. Gradients were foundto exist along petioles; they may have a greater resistanceto water flow than stolons. The relationship between water fluxand stem anatomy, and the importance of differential flow ratesthrough stolons and petioles to plant behaviour during waterstress, are discussed. Trifolium repens L., white clover, water relations     

Gibberellins and Inhibitors in Relation to Flowering of White Clovers (Trifolium repens L.)

Flower primordia appeared sooner and flowering intensity increasedas day length was extended beyond 8 h (short day) in two whiteclovers: cv. Tamar and cv. Tammisto, originating from Israeland Finland, respectively. Application of GA₃ under short dayscaused Tamar to flower more intensively as much as the doseof GA₃ was increased, but it did not affect the flowering ofTammisto. The level of gibberellin-like substances in stolon apices washigher under long (16 h) than under short days in both cultivars.The level of inhibitors extracted from leaves was not affectedby day length in Tamar. However, in Tammisto this level wasalmost twice as high under short than under long days. It issuggested that the presence of a relatively high level of inhibitorsin Tammisto when grown under short days interfered with theactivity of gibberellins in its effect on flowering. ABA wasamong the inhibitors: its application under long days had noeffect on flowering, but it increased the number of stolons.Possibly inhibitors other than ABA are involved in the preventionof flowering. Leaf expansion responded to day length and GA₃ and ABA applicationin a similar way as the response of flowering, i.e. towardsflowering the size of the leaves increased.