Effect of Sucrose on Starch Accumulation in and Adventitious Bud Formation on Embryos of Picea abies

Adventitious buds were initiated on embryos of Picea abies (L.)Karst. after a pulse treatment with cytokinin. The initial stagesof bud formation could take place on culture medium lackingsucrose, but sucrose was required for further development ofmeristematic centres into bud primordia and buds. Sucrose atone per cent was optimal for adventitious bud formation. Embryoscultured on media containing sucrose started to accumulate starchduring the first day. Starch accumulation occurred especiallyin the cortex cells where starch grains were frequently presentin the chloroplasts. The starch accumulation increased withhigher sucrose concentrations in the culture medium. Embryoscultured on medium lacking sucrose did not accumulate starchbefore the formation of meristematic centres. Starch accumulationwas never observed in meristematic cells from which adventitiousbud primordia developed. Picea abies (L.) Karst., Norway spruce, adventitious bud, starch accumulation, sucrose concentration     

Some Effects of Sucrose Concentration on Primordium Development in Excised Primary Roots

The effects of varying the amount of sucrose used to supplementthe culture medium maintaining the growth of excised roots ofPisum sativum L., Vicia faba L., Zea mays L. and Phaseolus vulgarisL., on the rates of primordium initiation and subsequent emergenceas lateral roots and on the duration of the interval betweenprimordium inception and emergence as a secondary root throughthe tissues of the primary have been investigated. Variation in the exogenous concentration of sucrose from 0.5to 8 per cent had little effect on the rate of primordium inceptionin Pisum and Vicia and the rates never reached the values obtainedfor the roots of the corresponding intact plants. Moreover,over the 6 day culture period lateral root emergence did notoccur in any of the excised roots of these two species. In contrast,each of the aspects of primordium development examined in theexcised roots of Zea and Phaseolus was markedly affected bythe amount of sucrose used to supplement the culture medium.In addition, in the presence of about 6 per cent sucrose, primordiumdevelopment in these cultured roots was very similar to thatin roots of the corresponding intact plants. The results indicate either that some factor necessary for primordiumdevelopment is present in adequate amounts in excised rootsof Zea and Phaseolus, but not in those of Pisum and Vicia, orthat the factors controlling such development are differentin the former and latter two species. Vicia faba L., Pisum sativum L., Zea mays L., Phaseolus vulgaris L., broad bean, garden pea, maize, dwarf bean, primordium development, sucrose concentration, cultured roots     

The Culture of Immature Pea Embryos

Pea embryos of a range of developmental stages were culturedin a defined medium in vitro for up to 16 days. The criticalfactor for successful culture was the osmotic pressure of themedium; for the stages studied this was provided by the incorporationof 18 per cent sucrose in the medium. The growth of embryosof a range of genotypes was compared; small seeded genotypescould grow at comparable rates in vitro to those attained invivo. The amount of protein synthesized in vitro was similarto that attained in vivo, whereas slightly higher and lowerlevels of starch and DNA respectively were attained in vitro.The roles of intrinsic and extrinsic factors in regulating embryogrowth were studied by comparing the growth in culture of embryosof different genotypes and of hybrid embryos derived from reciprocalcrosses. embryo culture, pea embryos, hybrid embryos, osmotic pressure     

Sucrose Transport in Isolated Immature Barley Embryos

Techniques have been devised to select immature barley embryosat various stages in their development, and to study their accumulationof sucrose in vitro. Isolated embryos accumulate sucrose overa period of several hours of which some 80 per cent is conservedas a pool of free sucrose and the remainder utilized in macromolecularsynthesis. The rate of sucrose uptake increases with embryodevelopment, however the specific activity of uptake remainsconstant, indicating that the transport processes are fullyoperative early in embryogenesis. From the kinetics of sucroseuptake it is deduced that facilitated transport predominatesat sucrose concentrations of 50 mM, while at higher concentrationspassive diffusion makes an increasing contribution to sucroseaccumulation. The substrate specificity and the sensitivityof sucrose transport to uncoupling agents, in addition to thestability of the pool of accumulated sucrose, are all indicativeof active transport playing a major role in the sucrose assimilationof developing barley embryos. Hordeum distichum (L.) Lam, barley, embryo, sucrose transport     

The Carbohydrate Nutrition of Tomato Roots: V. The Promotion and Inhibition of Excised Root Growth by Various Sugars and Sugar Alcohols

Iron is only consistently present in an available form in White'sroot culture medium if the inorganic salts are autoclaved withthe sugar. The substitution of ferric ethylenediamine-tetra-acetatefor the inorganic ferric salt of White's medium ensures ironavailability when the carbon source of the medium is renderedsterile by ether treatment and subsequently added to the remainderof the constituents which have been sterilized by autoclaving. The biological activity of sugars, and particularly of dextroseand laevulose, is altered by autoclaving them in presence ofthe inorganic salt solution of White's medium. The only sugar which supports a considerable growth of excisedtomato roots is sucrose. The activity of this sugar is not affectedby alcohol-precipitation nor is it dependent upon the simultaneouspresence of traces of its constituent mono-saccharides. Dextrose or laevulose or a mixture of the two sugars supporta low but sustained level of excised-root growth. All othersugars and sugar alcohols tested were inactive as carbon sources. The addition of sucrose at low concentration (0–2 percent.) to a medium containing dextrose as the main carbon compounddoes not make possible a level of growth comparable with thatobtained with an adequate sucrose supply. It has not been possibleto enhance the growth-rate of excised roots supplied with dextroseby previous presentation of this sugar under conditions permittingactive growth. Using media containing 'etherized' sucrose anddextrose, no evidence was obtained of any competitive inhibitionof sucrose utilization by dextrose. Certain sugars when added to a medium, containing the optimumconcentration of sucrose₁, markedly inhibited excised root growth.Thus l-sorbose, l- and d-arabinose, and d-xylose caused notless than 80 per cent, inhibition at a concentration of 0-5per cent. d-mannose and d-galactose completely inhibited growthat o-1 per cent. The oligosaccharides, dextrose, laevulose,and the sugar alcohols tested had, by contrast, very low inhibitoryactivity.     

Influence of Genotype and Environment on Growth of Barley Embryos in vitro

Isolated embryos of three contrasting barley genotypes werecultured in vitro on a range of media comprising 16 combinationsof sucrose (3–12 per cent) and 2, 4-D (0–8 mg 1^–1)concentration. Cultures were incubated at a range of temperaturesfrom 5 to 25°C and were examined after 21 days when shootlengths as well as fresh and dry weights were recorded. Therelative influence on growth of increasing concentrations ofsucrose and 2, 4-D was investigated, as was their interactionwith the incubation temperature. The genotypes were found todiffer markedly in their response to these two media components,with each parameter of growth differentially affected. Resultsare discussed in relation to the known dormancy characteristicsof these genotypes. Hordeum vulgare L., barley, embryo, dormancy, 2, 4-dichlorophenoxy acetic acid, sucrose     

Comparative Morphological Study of Zygotic and Microspore-derived Embryos ofBrassica napusL. as Revealed by Scanning Electron Microscopy

A comparative morphological study of microspore-derived (MD)and zygotic embryos ofBrassica napusL. was conducted, illustratingsubstantial similarities in external morphology of these embryosthroughout their development. Haploid embryos were producedfrom isolated microspores cultured on high molecular weightpolyethylene glycol (PEG), replacing sucrose as an osmoticum.Morphological changes during the time-course of microspore embryodevelopment induced on PEG (25%) and sucrose (13%) are describedin detail as revealed by scanning electron microscopy (SEM)and compared to the corresponding stages of zygotic embryosdevelopedin ovulo. At the heart, torpedo and early cotyledonarystages, microspore-derived (MD) embryos on PEG closely resembletheir zygotic counterparts. In contrast, the external morphologyof embryos induced on high sucrose medium differs from thatof PEG and zygotic embryos indicating that a high concentrationof sucrose in culture has a morphogenetic effect on MD embryodevelopment inB. napus. Fragments of the original pollen wallare regularly observed at the root pole region and at the tipsof suspensors in MD embryos throughout their development. Thissuggests that polarity in MD embryos might originate from structurallypolarized late uninuclear microspores and early bicellular pollen.Copyright1998 Annals of Botany Company Brassica napusL., scanning electron microscopy, microspore-derived embryo, zygotic embryo, morphology, microspore, suspensor, exine, sucrose, polyethylene glycol.     

Sulphate Transport into Plants and Excised Roots of Soybean (Glycine max L.)

The rates of sulphate transport into intact and excised rootsof soybean (Glycine max L.) were not significantly differentin the first hour and were maximal at pH 7. However, intactroots accumulated four times as much sulphate as excised rootsin 24 h, because of a marked reduction in the rate of transportby excised roots. The continued high rates of transport intointact roots were observed in plants kept in the light, andobserved in darkened plants growing in 1 per cent sucrose. Similarly,sulphate accumulation by excised roots was stimulated 2-foldby 1 per cent sucrose. The characteristics of sulphate accumulation by roots were notuseful in predicting sulphate translocation to the leaves. Transportto the leaves was maximal at pH 2–3, was almost totallylight-dependent and was not enhanced by growing plants in sucrose. Sulphate transport, Glycine max L., soybean, excised roots     

Expression of gus in somatic embryo cultures of black spruce after microprojectile bombardment

Immature embryos (stage I) and cotyledonary somatic embryos(stage III) of black spruce [Picea mariana (Mill) B.S.P.] werebombarded with tungsten particles coated with a gene constructcontaining the fusion of gus:: nptll. GUS (ß-glucuronidase)activity was monitored histochemically with X-gluc giving ablue colour where transient gene expression was detected inthe bombarded tissues. A high transient expression of gus wasobserved in stage I embryo cultures 2 d after bombardment (202GUS foci per 300 mg tissue). GUS activity had substantiallydiminished in this material 14 d after bombardment, when grownin liquid LP maintenance medium containing BA (4.4µM),2,4-D (9µM) and 1% sucrose. However, when stage I embryoswere cultured on LP maturation medium containing BA (40 µM),IBA (1 µM), 3.4% sucrose and 0.8% agar, GUS activity after2 d was 335 GUS foci per 300 mg tissue, and the activity wasdetected until 30 d after bombardment. With stage III somaticembryos cultured on LP maintenance medium, 92% showed GUS activity2d after bombardment (16 GUS foci per embryo), and 31 % showedactivity 30 d after bombardment (4 GUS foci per embryo). GUSactivity was still evident in 12% of the embryos (2 GUS fociper embryo) 45 d after bombardment. Key words: Black spruce, gus = E. coli geneuid A encoding ß-glucuronidase, nptll = gene encoding neomycin phos-photransferase, somatic embryos     

Sporulation in Rhizopus sexualis and Some Other Fungi Following a Period of Intense Respiration

Rhizopus sexualis grown at 20° C. on liquid I per cent.malt or glucose-asparagine medium showed a peak of respiratoryactivity between 40 and 55 hours-after inoculation. Rate ofrespiration then fell until it reached a steady low level whichcoincided with maximum mycelial growth. Zygospore initiationoccurred at or just after the peak of respiration. At a low temperature (9° C.) or with high concentrationsof glucose the respiration peak was less marked and no zygosporesdeveloped. Single ‘plus’ or ‘minus’strains of the heterothallic species Mucor hiemalis and Phycomycesblakesleeanus showed a pattern of respiration and mycelial growthsimilar to that of R. sexualis but no zygospores were formed.Zygospores did not develop without a preliminary period of intenserespiration, but such a peak period could occur without beingfollowed by zygospore formation. A strain of Sordaria fimicola was grown at 25° C. on a syntheticmedium with 5.0 per cent. sucrose or glucose as source of carbon.Respiration reached a peak at approximately 4 and 5 days respectively,the actual peak value being twice as large on sucrose as onglucose. Dry weight of mycelium was greater on glucose thanon sucrose. Perithecia were formed only on the sucrose medium.Visible peri-thecial initials were first seen shortly afterthe occurrence of the respiratory peak. Mature perithecia werepresent 3 days later. The possible significance of these results is discussed.     

Studies on the Growth and Respiration of Roots: I. THE EFFECTS OF STIMULATORY AND INHIBITORY CONCENTRATIONS OF {beta}-INDOLYLACETIC ACID ON ROOT SECTIONS OF PISUM SATIVUM

Simultaneous observations on extension growth and respirationrate (oxygen consumption) of 2-mm. sections excised from theextension zone of roots of pea (Pisum sativum) growing in distilledwater and 0·5 per cent. sucrose have yielded resultsclosely similar to those of Brown and Sutcliffe (1950). Respirationrate is not obviously correlated with growth rate either inwater or in sucrose, but it is strongly correlated with sectionlength. Respiration rate per unit section length (¬per unitfresh weight) shows a marked downward drift during extensionand is affected little by growth conditions. Tentative suggestionsare advanced to account for the small differences between driftsin o·5 per cent. sucrose and those in distilled water. Medium agitation produces an immediate and sustained stimulationof growth but no stimulation of oxygen uptake until the latergrowth stages. Thus respiration per unit section length is unaffectedby agitation at any stage. A typical growth response to ß-indolylacetic acid(IAA) was obtained, with a maximum stimulation (of about 35per cent.) at 1 part in 10¹¹ and inhibitions increasing progressivelywith concentration beyond the threshold of about i part in 10⁹.Both percentage stimulation and percentage inhibition of growthwere independent of the presence of sucrose. Respiratory responses to ß-indolylacetic acid werecomplex. In water no immediate response could be detected witheither a growth-stimulatory (10^–11) or a growth-inhibitory(10^–-8) concentration, while in 0·5 per cent. sucrosethe inhibitory concentration prevented the small immediate respiratoryrise due to the sucrose, probably by impeding sugar entry. Duringthe subsequent period of rapid growth (up to 36 hours) the smallrespiratory responses observed closely followed the small growthresponses to both concentrations of IAA, suggesting that theformer are the direct result of the differences in section lengthinduced by the auxin. When growth ceases (at 48 hours) sectionswhich have grown considerably in sucrose show respiratory ratesstill closely correlated with section length, whereas in waterboth concentrations of auxin induce marked depressions in respirationrate. It is concluded that ß-indolylacetic acid in bothgrowth-stimulatory and growth- inhibitory concentrations hasno direct effect on the activity of the respiratory enzyme systemof growing root cells. The small respiratory responses are bestexplained as resulting from differential changes in sectionsize and correlated changes in the enzyme complements of thegrowing cell.     

Establishment of in vitro Cultures of Tapinanthus bangwensis (Engler and K. Krause) Danser and the Effects of Sugars on Growth

Culturing of organs in vitro has been successfully employedin studies on morphogenesis and nutritional requirements ofparasitic and semi-paraaitic angiosperms. Tapinanthus bangwensis,a semi-parasite, has been successfully cultured on chemicallydefined media. By and large the parasite will thrive well ina medium of mineral salts and sucrose at its optimal concentration(4 per cent). However, the parasite is able to metabolize awide range of sugars most of which show similar concentrationoptima Although the growth in vivo was simulated in vitro inthe early stages, it was found that in the later stages growthin vitro was much slower than growth in vivo. The growth differencesobserved in the different media may reflect some of the physiologicaldifferences that are responsible for the selective nature ofthe parasite's development and establishment on different hosts     

Flavanoids in Pollen and Stigma of Brassica oleracea and Their Effects on Pollen Germination In Vitro

Brassica oleracea pollen was applied to a basic medium of 1.5per cent agar and 15 per cent sucrose to which flavanoids wereadded at three concentrations. Two types of agar were used;with agar 1, quercetin at a concentration of 0.5 x 10^–3per cent gave an increase in percentage germinating grains.With agar 2, an increase in germination occurred with kaempferoland naringin at concentrations of 0.5 x 10^–3 and 0.5 x10^–1 per cent respectively. Increase in pollen tube lengthoccurred with agar 2 and quercetin at a concentration of 0.5x 10^–3 per cent. The stigma tissue of B. oleracea contains at least three andthe pollen at least one glycoside of quercetin. The sugars inthe glycosides were not identified. Pollen germination and pollentube extension were not stimulated exclusively by the flavanoidspresent in the stigma. The flavanoid composition of the stigmadid not vary amongst five different S-allele genotypes, indicatingthat flavanoids are probably not directly involved in the incompatibilityreaction of B. oleracea.     

Storage Lipid Accumulation by Zygotic and Somatic Embryos in Culture

In vitro accumulation of storage lipids occurs in zygotic andsomatic embryos of Brassicu napus L. The concentration of sucrosein the medium modified the pattern of storage lipid accumulationin zygotic and somatic embryos. The sucrose concentration atwhich the maximum amount of lipid is accumulated by the twotypes of embryos is different Analysis of fatty acid compositionshowed that the same fatty acids are present in embryos in vivoand those cultured in vitro although there are quantitativedifferences. The possibility of using this type of system forin vitro production of valuable plant metabolites is discussed Embryo cloning, somatic embryogenesis, in cilro culture, storage lipids, Brussica napus, oilseed rape     

Mineral Ion Nutrition of Developing Barley Grains 1. Effect of Magnesium Deficiency on Ion Content of Immature Embryos and Endosperms

The effect on the mineral ion nutrition of developing barleyembryos and endosperms was studied when both Mg and Mn werewithdrawn simultancously from the rooting medium. Both embryosand endosperms had lower Mg levels, on average 40 and 50 percent, respectively, over the developmental period, than thosefrom control plants grown in complete nutrient solution. Comparedto the controls, higher levels of Mn (average 20 per cent) andCa (average 20 per cent) were found in Mg-deficient embryosfrom test plants, but Mn levels were lower (average 30 per cent)and Ca levels unaffected in Mg-deficient endosperms. K levelsin both embryos and endosperms were not changed. The endospermsfrom test plants had lower total N levels than the controls. Hordeum vulgare L., barley, embryo, endosperm, sand culture, magnesium, manganese, calcium, potassium, nitrogen, mineral nutrition     

The In Vitro Culture of Immature Barley Embryos on Different Culture Media

Immature barley embryos (Hordeum distichum var. Julia) of between0•20 and 0•80 mm in length, were isolated from thedeveloping grain and cultured in vitro on various culture media.The subsequent development of the embryos was followed overa period of weeks, and where germination ensued the growth rateof shoot and root meristems was compared with in vivo germinationrates. Various growth media were assessed for their abilityto support normal development of immature embryos. A numberof published media failed to support satisfactory developmentof young embryos. The addition of 1–15 per cent coconutmilk to Norstog's Medium I (mineral + vitamin solns) enhancedembryo development and lowered the threshold of viability fromembryos of 0•50 mm in length to 0•35 mm. Althoughin many cases germination ensued, embryo development was largelyabnormal. A slightly greater enhancement of growth was achievedwith 0•05–0•30 per cent casein hydrolysate asthe growth medium supplement, although abnormal developmentwas not eliminated. A further lowering of the viability thresholdto include embryos of 0•25 mm in length was obtained bycombining 2•7 mM glutamine with the casein hydrolysatesupplement. Normal development and germination of embryos assmall as O25 mm was however obtained on Norstog's Medium JJand the results were reproduced in four additional if . distichumvarieties. In each case the critical threshold of viabilitywas found to lie in embryos of 0•20–0•30 mmin length.     

Lateral Root Anlage Development in Excised Roots of Vicia faba L., Pisum sativum L., Zea mays L. and Phaseolus vulgaris L.

The development of lateral root primordia has been investigatedin excised roots of Vicia faba, Pisum sativum, Zea mays andPhaseolus vulgaris cultured in White's medium supplemented with2 per cent sucrose and compared with previously published dataon such development in primaries of the corresponding intactplants (control roots). Primordia were produced in each batchof excised roots over the 6 day culture period but at a lowerrate (number day^–1) than in the controls. Such primordia in cultured roots of Zea and Phaseolus completedtheir development and grew out as lateral roots over a periodsimilar in length to that found in the controls, but with acell number of only about 33 per cent of that attained at thetime of secondary emergence in the primaries of the latter roots.These lower cell numbers were at least partly a reflection ofincreases in mean cell doubling time over the period of anlagedevelopment investigated in the excised roots relative to thecorresponding values found in the controls. Primordia initiated in excised roots of Pisum and Vicia didnot complete their development in culture, i.e. no lateral rootsemerged and arrest took place with cell numbers of only 37 (Pisum)and 17 (Vicia) per cent of the numbers determined at the timeof secondary root emergence in the controls. Such arrested primordiahad few nuclei in S and none in mitosis. Moreover, at leastin Pisum, the frequency distribution of the relative DNA contentof the nuclei in the latter primordia approximated that foundin the apical meristem of primary roots following the establishmentof the stationary phase under conditions of carbohydrate starvation. It has also been demonstrated in the course of these investigationsthat lateral root primordium development in all four speciesis at least biphasic and possibly triphasic. Vicia faba L., broad bean, Pisum sativum L., garden pea, Zea mays L., maize, Phaseolus vulgaris L., dwarf bean, root primordia, anlage, cell doubling time, lateral root emergence     

Effect of Post-anthesis Drought on Cell Division and Starch Accumulation in Developing Wheat Grains

Wheat plants (Triticum aestivum L., cv. Warigal) were subjectedto 20 d of water deficit during the period of endosperm celldivision. Drought accentuated the differences in final grainweight between spikelets and between grains within spikelets.The distal grains of top spikelets were most affected by drought.The maximum number of endosperm cells was, respectively, 30and 40 per cent lower in basal grains and distal grains of draughtedplants. In basal grains of middle spikelets, the number of largestarch granules per cell was unaffected but the number of smallstarch granules per cell was 45 per cent lower in grains ofdraughted plants. The initiation of small starch granules wasmore affected than cell division because severe water deficitoccurred earlier during the former process than the latter.Final dry weight appeared to correlate well with the maximumnumber of endosperm cells, but depended also on the number ofstarch granules per cell. Consequently, the amount of dry matterper cell was not constant in both treatments. The concentration of sucrose per endosperm cell was lower onlyin the droughted distal grains of top spikelets. The supplyof sucrose to endosperm cells did not regulate the initiationof small starch granules. Triticum aestivum L., wheat, drought, grain growth, cell division, starch     

Microbiological Synthesis of Fat: THE FORMATION OF FAT FROM SUCROSE

1. Forty-three strains of moulds, selected from ten species,were examined to ascertain their capabilities as producers offat, when grown in four different solutions of nutrient inorganicsalts together with sucrose. The three highest yields of feltwere given by Penicillium javanicum van Beyma, P. Soppi Zaleski,and Aspergillus nidulans Eidam, in that order. The fat contenton felt weight was maximal at 34·8 per cent, with P.Soppi, at 28·4 per cent. with Fusarium lini(1), and at25·8 per cent. with A. nidulans; on sugar utilized itwas maximal at 11·4 per cent. with P. Soppi, at 7·9per cent, with A. nidulans, and at 5·6 per cent. withF. lini(1). 2. Other moulds considered worthy of further trial were P. spinulosum,P. aurantiobrunneum, F. graminearum, A. flavus, F. oxysporum,A. clavatus, and P. oxalicum.     

Soya Bean Seed Growth and Maturation In vitro without Pods

Immature Glycine max (L.) Merrill seeds, initially between 50and 450 mg f. wt, were grown and matured successfully in vitro.Excised seeds were floated in a liquid medium containing 5 percent sucrose, minerals and glutamine in flasks incubated at25 °C under 300 to 350 µE m^–2 s^–1 fluorescentlight. During 16 to 21 d in culture, seeds grew to a matured. wt of 100 to 600 mg per seed at an average rate of 5 to 25mg d. wt per seed d^–1 depending on initial size. Growthrates were maximal during the first 8 to 10 d in vitro but declinedwith loss of green colour in the cotyledons. Seed coats rupturedwith rapid cotyledon expansion during the first 2 d in culture.Embryos were tolerant to desiccation and 80 to 90 per cent germinatedif removed from culture before complete loss of green colour.The growth of excised seeds in vitro exceeded the growth ofseeds in detached pods, but when windows were cut in pods topermit direct exposure of seeds to the medium, seed growth wascomparable. Glycine max (L.) Merrill, soya bean, seed culture, seed growth, seed maturation, germination