Dynamics of extractable carbohydrates in Pisum sativum. II. Carbohydrate content and photosynthesis of pea cuttings in relation to irradiance and stock plant temperature and genotype

Rooting ability was studied for cuttings derived from stock plants of wild type pea seedlings and seedings of two mutants deficient in photosystem II activity and chlorophyll. Stock plants were grown at 15, 20, 25 or 30°C at 38 W m^-2. Cuttings were rooted at 20°C and at an irradiance of 16 or 38 W m^-2. The rooting ability seemed to be correlated with the initial carbohydrate content only at 38 W m^-2. Based on the findings of the present study it may be concluded that for pea seedlings the growth temperature is more important than photosynthesis as regards accumulation of extractable carbohydrates. During the rooting period carbohydrates are necessary for root formation, but the effect of the iradiance on the number of roots formed is not mediated by the carbohydrate content. Under specific rooting conditions it is possible to correlate the initial carbohydrate content with the rooting capacity of the cuttings within a phenotype, but not always when different phenotypes are considered. The results indicate a connection between the metabolic activity of the cuttings and their ability to form adventitious roots.     

Dynamics of extractable carbohydrates in Pisum sativum. I. Carbohydrate and nitrogen content in pea plants and cuttings grown at two different irradiances

Two methods of fixation of ³H-IAA on macromolecules were studied in order to obtain autoradiographs of semi-thin sections after a routine treatment for electron microscopy. Glutaraldehyde and DCC [1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride] could be used to obtain in vitro fixation of auxin on proteins. However, the rate of reaction with DCC was greater, so that the method using this compound is preferable. A cytological study was performed on wheat coleoptiles ( Triticum sativum L., var. Capitole) after application of ³H-IAA to their tips. Almost no radioactivity was found in the phloem elements while the procambium and the vessels were intensively labelled. In parenchyma cells, cell walls, nuclei and intercellular spaces contained a large amount of ³H-IAA. These results are consistent with the hypothesis that auxin is transported, from cell to cell, via the apoplast. The extraction of auxin by methanol before cytological fixation indicates that this hormone is not bound (by a covalent binding) to cellular structures, except perhaps in the secondary wall of vessels.     

A relationship between irradiation, carbohydrates and rooting in cuttings of Pisum sativum

Rooting ability was studied for cuttings derived from pea plants ( Pisum sativum , L. cv. Alaska) grown in controlled environment rooms. When the cuttings were rooted at 70 μmol m⁻² s⁻, ¹ (photosynthetic photon flux density) or more, a stock plant irradiance at 100 μmol m⁻² s⁻¹ decreased rooting ability in cuttings compared to 5 μmol m⁻², s⁻¹, However, cuttings rooted at 160 μmol m⁻² s⁻¹ formed more roots compared to 5 (μmol m⁻² s⁻¹. Although a high irradiance increased the number of roots formed, it could not overcome a decreased potential for root formation in stock plants grown at high irradiance. Light compensation point and dark respiration of cuttings decreased by 70% during the rooting period, and the final levels were strongly influenced by the irradiance to the cuttings. Respiratory O₂ uptake decreased in the apex and the base of the cutting from day 2 onwards, whereas a constant level was found in the leaves. Only the content of extractable fructose, glucose, sucrose and starch varied during the early part of the rooting period. We conclude that the observed changes in the cuttings are initiated by excision of the root system, and are not involved in the initiation of adventitious roots.     

Interaction of gibberellic and indole-3-acetic acid on root formation in pea (Pisum sativum L.) epicotyl cuttings

Indole-3-acetic acid (IAA) strongly enhanced rooting of etiolated pea epicotyl cuttings while gibberellic acid (GA₃) enhanced rooting only slightly. The promoting effects of the hormones appeared not until 14 d after the onset of treatment. When GA₃ and IAA were applied together, the initiation of rooting started already after 6 d after onset of treatment. It is suggested that gibberellin plays an important role, in combination with auxin, in the initiation of root formation in Pisum cuttings.Abbreviations IAA Indole-3-acetic acid - GA₃ Gibberellic acid     

Induction of somatic embryos in pea,Pisum sativum L.

Using low concentrations of picloram (0.06 mg/l), embryoids were formed on the surface of leaf-derived callus of pea, Pisum sativum L. (c.v. Dippes Gelbe Victoria) upon transfer to liquid medium. After some days in culture, embryoids spontaneously separated from the calli, and developed into torpedo-shaped embryos, which were transferred to solid medium. In a second series of experiments, embryos were also formed by mutant 489C and a genetic line of Pisum arvense, which additionally exhibited embryogenesis also from epicotyl-derived callus. Some of the embryos showed root formation, but no shoot morphogenesis occurred. In a limited number of cases, an additional root was formed in the apparent shoot apical region after 2–5 days.     

Synergistic effect of gibberellic acid and indole-3-acetic acid on rooting in stem cuttings of Abelmoschus esculentus Moench

Indole-3-acetic acid (IAA) and gibberellic acid (GA₃) enhanced the formation of roots on the stem cuttings of Abelmoschus esculentus. The effect increased considerably when both IAA and GA₃ were applied together.     

Physical conditions of propagation media and their influence on the rooting of cuttings

Summary The formation and subsequent growth of roots by cuttings of poinsettia, hydrangea, rose and azalea in various propagation media, Jiffy-7, Jiffy-9 and Grodan under different conditions of aeration was investigated. The interrelationships of the effects of air content of the media, temperature and light intensity on the rooting of poinsettia cuttings was also studied.With low air contents (0 cm moisture tension) in the propagation media the formation and growth of roots was strongly inhibited. The rooting performance of rose appeared to be less affected by the poor aeration. Increasing air content improved rooting but best results were obtained at moisture tensions of 4 to 8 cm. Rooting seems to be better correlated with oxygen diffusion rate (ODR) than with air content.For poinsettia cuttings the optimum temperature for rooting was 24 to 28°C. At low temperatures rooting was delayed while at higher temperatures it was almost completely inhibited. Callus formation increased with temperature but decreased with increasing moisture tension. Conditions which induced large callus formation inhibited root formation.High light intensity during rooting reduced overall rooting performance and the inhibition was most pronounced in conjunction with high moisture tensions.Report No. 255.     

The effect of mechanical impedance on root growth in pea (Pisum sativum). II. Cell expansion and wall rheology during recovery

The aim of the present work was to determine the factors limiting growth in mechanically impeded roots. Pea roots were grown in compressed and uncompressed sand cores, and then removed and transferred to hydroponics. Root elongation was slowed in impeded sand cores and did not recover to the unimpeded rates until 60 h after transfer to the hydroponics system. Root diameter was greater in impeded roots, and only after 36 h in hydroponics was new root tissue produced of the same diameter as the unimpeded controls. The turgor pressure of the growing cells was measured with a turgor probe and was the same in both treatments. The slower elongation rate of the previously impeded roots was, therefore, the result of axial tightening of the cell walls. Cell length profiles suggested that axial cell wall tightening persisted in the unrestricted hydroponics system. Production of new cells in unrestricted conditions was required before root elongation returned to the unimpeded state. Osmotic potential was decreased by approximately 0.2 MPa in previously impeded roots compared with the unimpeded ones. This corresponds to a decrease in water potential of 0.2 MPa. These data are discussed in relation to regulation of cell extension, solute unloading and the penetration of compacted soils by roots.     

Purification and properties of nitrite reductase from roots of pea (Pisum sativum cv. Meteor)

Nitrite reductase (EC 1.6.6.4) prepared from pea roots was found to be immunologically indistinguishable from pea leaf nitrite reductase. Comparisons of the pea root enzyme with nitrite reductase from leaf sources showed a close similarity in inhibition properties, light absorption spectrum, and electron paramagnetic resonance signals. The resemblances indicate that the root nitrite reductase is a sirohaem enzyme and that it functions in the same manner as the leaf enzyme in spite of the difference in reductant supply implicit in its location in a non-photosynthetic tissue.Abbreviations DEAE diethylaminoethyl - EPR electron paramagnetic resonance - NIR nitrite reductase - SDS-PAGE sodium dodecyl sulphate-polyacrylamide gel electrophoresis     

The effect of the duration of callus culture on the accumulation of genetic alterations in pea Pisum sativum L.

Two groups of regenerant plants were obtained from different pea genotypes (lines R-9 and W-1 and cultivar Viola). The first group was derived after eight months of culture and the second, from calluses cultured for a prolonged (more than ten years) time. Using random amplified polymorphic DNA (RAPD) and inter simple sequence repeats (ISSR) methods, the regenerants and the original lines were compared with regard to genetic differences. The regenerants from both groups were shown to differ in DNA polymorphism from the original lines and from one another. The divergence of the regenerants was also different, depending largely on the original genotype. Examination of genetic differences between the first and the second group showed that the variability increased with culturing time. This was particularly evident for regenerants of the Viola cultivar, in which variability ranged from 0–5% (first group of regenerants) to 10% (second group of regenerants).     

Development of the quiescent center in maturing embryonic radicles of pea (Pisum sativum L. cv. Alaska)

Maturing embryos of pea (Pisum sativum L. cv. Alaska) were treated with an aqueous solution of tritiated thymidine for 1 h, sectioned, and processed for autoradiography. An analysis of the distribution of labelled nuclei and mitotic figures demonstrated the presence of a quiescent center (QC) in the radicles of developing embryos. The QC developed in the radicle during the growth of the embryo. Immature radicles that did not contain a well-formed zone of root-cap initials did not show a QC. In the latter stages of seed ripening, the pattern of arrest of DNA synthesis and mitosis was tissue-specific. Cells within the QC remained inactive. The region lacking labelled nuclei and mitotic figures progressively expanded to include the root cap initials and then the provascular cylinder. Mitosis was arrested before DNA synthesis in the embryonic cortex. Cells within the QC synthesized DNA during the first stages of seed germination.Abbreviations [³H]TdR tritiated thymidine - QC quiescent center     

Reprogramming of cells following wounding in pea (Pisum sativum L.) roots. II. The effects of caffeine and colchicine on the development of new vascular elements

Summary Interference with the normal progression of the cell cycle by the drugs caffeine and colchicine does not prevent parenchyma cells in the cortex of pea roots from being reprogrammed to become tracheary and sieve elements following severance of the vascular cylinder of the root. The pattern of secondary wall deposition of the newly differentiated tracheary elements is highly aberrant in the presence of colchicine but is of normal appearance in the caffeinetreated roots. In each case, the new sieve elements have sieve plates and lateral sieve areas with callose deposits. Induction and redifferentiation are achieved in the absence of cell division and microtubules in colchicine-treated roots.Bi- and multi-nucleate cells are produced by both drugs. Microtubules are still present in the caffeine-treated roots but cell plate formation is inhibited. The partitioning of the multinucleate cortical cells by the interdigitation of free-growing walls between the nuclei occurs in the presence of caffeine but not colchicine.     

The effect of surface soil consolidation on the early root development of pea (Pisum sativum L.)

Cores of repacked soil were consolidated with a compressive strength testing machine, after peas had been planted in the centre of the core. The number that emerged were counted and root and shoot lengths and diameters were measured. Consolidation had no effect on emergence, root length or root diameter of the peas grown in a loamy sand, whereas emergence, root length and root diameter were affected by a small increase in load in a clay loam.     

The effect of mechanical impedance on root growth in pea (Pisum sativum). I. Rates of cell flux, mitosis, and strain during recovery

We studied the effect of mechanical impedance on cell flux and meristematic activity in pea roots. Pea seedlings ( Pisum sativum L. cv. Helka) were grown in cores of sand packed to dry bulk densities of either; 1.4 Mg m⁻³ with an additional 2.4 kg uniaxial load applied to the surface to increase the mechanical resistance to growth (penetration resistance of 1.5 MPa); or 1.0 Mg m⁻³ (penetration resistance of 0.05 MPa). A water content of 0.06 g g⁻¹ was chosen for optimum root growth. After 3 days, the seedlings were transferred to hydroponics, colchicine was added and the rate of cell doubling, mitotic index and length of the cell cycle was assessed. Cell flux in the third cortical layer was calculated for roots immediately removed from sand.Mechanical impedance slowed root extension to about 20% of the unimpeded rate, and final cell length was reduced to 50% of the unimpeded length. The rate of cell doubling was 3.4 times slower for roots recovering from mechanical impedance mostly as a result of a longer period spent in interphase. Cell flux in impeded roots was approximately half that of unimpeded roots (5 cells h⁻¹), and contributed to a shorter cell file and elongation zone, and a slower rate of root elongation.     

The Effect of Triacontanol on Peroxidase, IAA, and Plant Growth in Pisum sativum var.'Alaska' and 'Little Marvel'

The present study investigates the effects of triacontanol (CH₃(CH₂)₂₈CH₂OH),on plant growth (root and stem), peroxidase activity (apicalmeristem tissue), and auxin destruction (apical meristem tissue)in ‘Little Marvel’ dwarf (LM) and ‘Alaska’peas (AP). Triacontanol inhibited root growth in LM comparedto untreated controls. However, root growth in AP tissue wasenhanced by 1.0 mg I^–1 triacontanol and inhibited by allother treatments, in comparison to untreated controls. Wateruptake in triacontanol-treated AP plants was greater than inuntreated controls, with the converse being the case for LM.Triacontanol treatment caused an increase in peroxidase activityin both LM and AP plants compared to untreated controls. Interms of (1–¹⁴C)IAA destruction, GA₃ + 0.01 mg 1^–1triacontanol caused appreciable auxin breakdown (40%) in LMtissue, with GA₃ + 0.1 mg 1^–1 triacontanol giving a 43%decrease compared to untreated controls. In AP tissue, 10 µMGA3 increased auxin destruction by 188% whereas 0.1 mg I^–1triacontanol caused a 20% decrease compared to untreated controls.The effects of triacontanol on root and stem growth, peroxidaseactivity, and auxin destruction appear to be cultivar-specific,with respect to LM and AP varieties of peas.     

The interaction of xylosyltransferase and glucuronyltransferase involved in glucuronoxylan synthesis in pea (Pisum sativum) epicotyls.

A particulate enzyme preparation from etiolated pea (Pisum sativum) epicotyls was found to incorporate xylose from UDP-D-xylose into beta-(1----4)-xylan. The ability of this xylan to act as an acceptor for incorporation of [14C]glucuronic acid from UDP-D-[14C]glucuronic acid in a subsequent incubation was very limited, even though glucuronic acid incorporation was greatly prolonged when UDP-D-xylose was present in the same incubation as UDP-D-[14C]glucuronic acid. This indicated that glucuronic acid could not be added to preformed xylan. However, the presence of UDP-D-glucuronic acid inhibited incorporation of [14C]xylose from UDP-D-[14C]xylose into beta-(1----4)-xylan, and neither S-adenosylmethionine nor acetyl-CoA stimulated either the xylosyltransferase or the glucuronyltransferase.     

Effects of cytokinin on ethylene production and nodulation in pea (Pisum sativum) cv. Sparkle

In this study, we were interested in learning if cytokinins play a role in the developmental process that leads to nodulation in the pea cv. Sparkle. We demonstrate that the application of the synthetic cytokinin BAP (6-benzyl-amino-purine) results in a number of nodulation-related changes. BAP stimulates the production of ethylene, a known inhibitor of nodulation. At low levels (up to 1 μ M ), BAP also stimulates nodulation but as its concentration is increased (up to 25 μ M ), nodule number decreases. In BAP-treated roots, the infection threads are abnormal; they are twisted, very knotty, and generally grow in a direction parallel to the root surface. In addition, the centers of cell division in the inner cortex are very few. Thus, BAP-treated Sparkle appears to phenocopy the low-nodulating pea mutant R50 [Guinel FC, Sloetjes LL (2000) Ethylene is involved in the nodulation phenotype of Pisum sativum R50 ( sym 16 ), a pleiotropic mutant that nodulates poorly and has pale green leaves. J Exp Bot 51: 885–894]. However, it appears doubtful that there is a direct correlation between the actions of cytokinin and ethylene in causing a reduction in nodule organogenesis because nodulation is not restored by treating BAP-treated Sparkle with ethylene inhibitors.     

The effects of sodium chloride-salinity upon growth, nodulation, and root nodule structure of pea (Pisum sativum L.) plants

Pea (Pisum sativum L.) plants inoculated with Rhizobium leguminosarum bv. viciae effective strain 248 were irrigated with nitrogen-free medium supplemented with 0, 25, 50 or 75 mM NaCl. The inhibitory effect of salinity on the growth of pea plants treated with 25 mM NaCl occurred 6 weeks post inoculation. In case of 75 mM NaCl treatment, the same effect was observed 2 weeks post inoculation. In contrast to investigations described in the literature our results clearly indicated that 25 mM NaCl stimulated nodule formation, however, in contrast to control nodules (the medium without NaCl), the nodules were considerably smaller. Remaining variants of salt treatment reduced plant growth, nodulation, and total nodule volume calculated per plant. Microscopic observations showed that salinity: (1) caused the loss of turgor of the nodule peripheral cells, (2) changed nodule zonation, (3) stimulated infection thread enlargement and expansion, (4) caused disturbances in bacterial release from the infection threads, and (5) induced synthesis of electron dense material (EDM) and its deposition in vacuoles. It was also found that cisternae of RER were involved in the formation of special cytoplasmic compartments responsible for synthesis of EDM. Autofluorescence study revealed that salinity increased accumulation of phenolics in pea nodules, as well.