Ethylene and cytokinin interaction in the morphogenesis of Pelargonium × hortorum L.H. Bailey in vitro

Pelargonium × hortorum ‘Grand Prix’ which is susceptible to leaf yellowing and ‘Bergpalais’ which is not susceptible to leaf yellowing were chosen for the experiments. Ethylene production and action as well as the associated morphological response of Pelargonium shoots grown in the presence of a precursor of ethylene biosynthesis 1-aminocyclopropane-1-carboxylic acid (ACC), ethylene inhibitors: α-aminooxyacetic acid (AOA) and silver nitrate (AgNO₃) and different cytokinins: (meta-topolin) (mT) or 6-benzylaminopurine (BAP) were studied. It was found that ‘Grand Prix’ was more sensitive to ethylene than ‘Bergpalais’ and it showed the leaf yellowing in response to 0.1 mg l^?1 ACC. Moreover, it was noted that ACC added separately or together with cytokinin influenced Pelargonium morphogenesis. Depending on the concentration of ACC (0.1–2.0 mg l^?1), it either stimulated or inhibited shoot and root formation as well as the growth of shoots and leaf blades. ACC-induced leaf yellowing in ‘Grand Prix’ was effectively inhibited by mT. In contrast, BAP did not enhance shoot quality. Simultaneously, the presence of mT in the medium resulted in up to a twofold increase in the ethylene production by ‘Grand Prix’ shoots throughout the culture period compared with the shoots growing on the BAP-medium. The inhibitor of ethylene action (AgNO₃) added with cytokinin prevented the yellowing of Pelargonium shoots, but simultaneously influenced the formation of mature shoots with limited long-term multiplication potential. The shoots of P. × hortorum ‘Grand Prix’ treated with AgNO₃ and mT emitted two- and sevenfold more ethylene after 11th and 21st day of culture compared with those treated with AgNO₃ and BAP. It is suggested that mT inhibits the early senescence of Pelargonium in vitro by decreasing its sensitivity to ethylene.     

Effect of growth regulators on maturation of geranium (Pelargonium × hortorum) somatic embryos

Interactions of growth regulators and polyethylene glycol on maturation of geranium somatic embryos were investigated. Somatic embryos were induced on medium with 20 M thidiazuron for 3 days. The growth regulators used were 1 µM abscisic acid, jasmonic acid, napthaleneacetic acid and benzylaminopurine at 21 days from the start of induction. Benzylaminopurine and napthaleneacetic acid did not enhance abscisic acid effects on maturation frequency but only improved maturation frequency in the presence of polyethylene glycol. Abscisic acid significantly improved protein content in the presence of polyethylene glycol. Benzylaminopurine and napthalene acetic acid in combination with abscisic acid and jasmonic acid improved protein types in somatic embryos only in the absence of polyethylene glycol. Osmoticum effected by polyethylene glycol seems the main component required for protein synthesis. This study showed significant improvement of somatic embryo quality for artificial seed production.     

Molecular fate of thidiazuron and its effects on auxin transport in hypocotyls tissues of Pelargonium × hortorum Bailey

Thidiazuron, a synthetic phenylurea-type cytokinin, has previously beenfound to induce somatic embryogenesis and organogenesis in a wide range ofplantspecies and to modulate the metabolism of endogenous auxins and cytokinins. Inspite of these findings, the precise mode of action of TDZ remainsundetermined.The current studies were undertaken to determine the fate of the TDZ moleculeand the effects of TDZ exposure on auxin transport in plants. The fate of tworadiolabelled versions of thidiazuron, [¹⁴C-5-thidiazol]-TDZ and[¹⁴C-U-phenyl]-TDZ, was investigated in sterile hypocotyl culturesofgeranium (Pelargonium×hortorumBailey). Radiolabelled TDZ was recovered from the tissue explants inethanol-insoluble, ethanol-soluble and chloroform fractions as well as inacidic, basic and neutral eluants from Dowex resins. Hypocotyl sections thathadbeen exposed to TDZ were found to accumulate more ¹⁴C-IAA from theculture medium and to translocate the auxin over a greater distance within thetissues. These data provide the first evidence that the TDZ molecule remainsintact in both a free and conjugated form within the plant tissues and providesome indication that TDZ-exposure enhances the accumulation and translocationofauxin within the tissues.     

Does ethylene play a role in thidiazuron-regulated somatic embryogenesis of geranium (Pelargonium × hortorum bailey) hypocotyl cultures?

Summary The accumulation of ethylene in headspace of hypocotyl cultures of geranium (Pelargonium × hortorum Bailey) and its possible role in thidiazuron-mediated somatic embryogenesis was investigated. The action of ethylene as determined by various ethylene synthesis and action inhibitors was varied. Silver nitrate (AgNo₃), aminoethoxyvinylglycine (AVG), and silver thiosulphate (STS) had no significant influence on the embryogenic response, while 1-methylcyclopropene (1-MCP) applied during the initial 3 d of induction or the expression phase, significantly increased the number of somatic embryos formed. Thidiazuron-treated tissues accumulated large quantities of ethylene within 6 h of culture, but the levels decreased after 12 h and reached very low levels after 3 d in culture. In the presence of acetylsalicylic acid (ASA), the levels of ethylene decreased by 20 to 50% during the first 48 h of culture. Analysis of endogenous auxin, cytokinins, and abscisic acid (ABA) indicated possible interactions of ethylene with other phytohormones during the induction of somatic embryos on geranium hypocotyl explants. Thidiazuron (10 μM) increased, while ASA decreased the levels of endogenous auxin, cytokinins, and abscisic acid during this period of induction.     

Effect of picloram and methyl jasmonate on growth and taxane accumulation in callus culture of Taxus × media var. Hatfieldii

We have analysed the effect of some culture conditions and media components on callus growth rate and production of taxanes in callus of Taxus × media var. Hatfieldii. For callus induction and maintenance a Gamborg B5 medium and a White - Rangaswamy medium (WR) with different modifications were used. On an improved WR medium (containing 10 μM picloram) the callus growth factor increased up to 5.8 fold (fresh weight). Picloram only enhanced the growth of callus, but not taxane production. On WR medium with (100 μM) methyl jasmonate the paclitaxel content increased from 2.37 μg g-1 to 90 μg g-1 and cephalomannine from 5.14 μg g-1 to 29.14 μg g-1 (dry weight), whereas growth of the cultures ceased. The presence of paclitaxel and cephalomannine was established by high performance liquid chromatography. This revised version was published online in June 2006 with corrections to the Cover Date.     

Effect of tropospheric ozone on morphological characteristics of Nicotiana tabacum L., Phaseolus vulgaris L. and Petunia×Hybrida L. in ambient air conditions

The cumulative ozone effect on morphological parameters (visible leaf injury, plant height and leaf growth, number of bean pods, petunia flowers and stalks) was examined in this study. Well-known ozonesensitive (Bel W3) and ozone-resistant (Bel B) tobacco cultivars as well as bean cv. Nerina and petunia cv. White cascade, both recognized as ozone sensitive, were used in the experiment. Investigations were carried out at two exposure sites varying in tropospheric ozone levels. Ozone negatively affected the leaf growth of both tobacco cultivars and bean. A negative relation was also found for ozone concentration and tobacco plant height. Number of petunia flowers and stalks and bean pods was positively correlated with ozone concentration. This could have been connected with earlier plant maturation due to faster generative development of plants in ozone-stress conditions.     

Effect of chitosan on tobacco mosaic virus（TMV） accumulation,hydrolase activity,and morphological abnormalities of the viral particles in leaves of N. tabacum L. cv. Samsun

The effect of chitosan on the development of infection caused by Tobacco mosaic virus（TMV） in leaves of Nicotiana tabacum L. cv. Samsun has been studied. It was shown that the infectivity and viral coat protein content in leaves inoculated with a mixture of TMV（2 μg/mL） and chitosan（1 mg/mL） were lower in the early period of infection（3 days after inoculation）, by 63% and 66% respectively, than in leaves inoculated with TMV only. Treatment of leaves with chitosan 24 h before inoculation with TMV also caused the antiviral effects, but these were less apparent than when the virus and polysaccharide were applied simultaneously. The inhibitory effects of the agent decreased as the infection progressed. Inoculation of leaves with TMV together with chitosan considerably enhanced the activity of hydrolases（proteases, RNases） in the leaves, in comparison with leaves inoculated with TMV alone. Electron microscope assays of phosphotungstic acid（PTA）-stained suspensions from infected tobacco leaves showed that, in addition to the normal TMV particles（18 nm in diameter, 300 nm long）, these suspensions contained abnormal（swollen, “thin” and “short”） virions. The highest number of abnormal virions was found in suspensions from leaves inoculated with a mixture of TMV and chitosan. Immuno-electron microscopy showed that “thin” virus particles, in contrast to the particles of normal diameter, lost the ability to bind to specific antiserum. It seems that the chitosan-induced activation of hydrolases stimulates the intracellular degradation of TMV particles and hence hydrolase activation may be considered to be one of the polysaccharide-mediated cellular defense mechanisms that limit virus accumulation in cells.     

Effect of soil cadmium application and pH on growth and cadmium accumulation in roots, leaves and fruit of strawberry plants (Fragaria × ananassa Duch.)

Three strawberry (Fragaria × ananassa Duch.) cultivars Rainier, Totem and Selva were grown under greenhouse conditions in a Parkhill sandy loam soil with a background DTPA-extractable Cd concentration of 0.18 mg kg^-1 and a pH of 5.1. Experimental treatments included combinations of 4 Cd applications (0, 15, 30 and 60 mg Cd kg^-1 soil) applied as CdSO₄ and 2 soil pH values 5.1 and 6.8. Both the application of Cd and pH of the soil significantly affected plant growth, yield and Cd accumulation in plant tissue anf fruit. Although roots accumulated the highest concentrations of Cd of all plant parts investigated, increased soil Cd application reduced leaf weight more than root weight. In general, yield of strawberries was decreased by an increase in amount of soil-applied Cd, however the yield response varied among cultivars. At 60 mg Cd kg^-1 soil, yield of Rainier cultivar was reduced to 17.6% of control plants. Over 90% of total Cd taken up by plants grown in Cd-treated soil accumulated in roots, regardless of the Cd level in the soil. Root Cd concentrations ranged from 2.6 mg kg^-1 (control plants) to 505.7 mg kg^-1 (Totem plants grown in soil at highest Cd and a soil pH 5.1) and were directly related to soil Cd concentrations. Cd translocation from roots to leaves and fruit was very limited, resulting in a maximum Cd concentration in root leaf tissue of 10.2 mg kg^-1. Accumulation of Cd in fruit was found to correlate well with leaf Cd, although even at the highest amount of applied Cd, fruit Cd concentration did not exceed 700 g kg^-1 of fresh weight.Contribution no. 951     

Effect of Ca/Mg ions ratio on copper accumulation, photosynthetic activity and growth of Cu2+-treated Salix viminalis L. ‘Cannabina’

The aim of the study was to the assess the influence of Ca/Mg ions ratio on the photosynthetic activity of Salix viminalis L. ??Cannabina?? plants cultivated in medium enriched with Cu(NO₃)₂. The experiment was conducted in controlled conditions in a phytotron for 21 days; hence the early plant response was tested. Plants were cultivated with different Ca/Mg ions ratios, i.e. (4:1)_l, (4:1)_h, and 1:10. Plants were additionally treated with Cu(NO₃)₂ at 1, 2, and 3 mM concentration in cultivation medium. Net photosynthetic rate, stomatal conductance and transpiration were measured after the first, second and third week of cultivation. Additionally, chlorophyll content, leaf morphology, root biomass and copper accumulation in leaves and roots were investigated. The investigations revealed differences in plant response to particular treatments ?? differences in Cu accumulation for particular Ca/Mg ions ratios were detected. It seems that plants are adapted to high Cu²⁺ concentrations, when 1:10 Ca/Mg ions ratio is applied. The highest Cu accumulation in roots was noted for plants fertilized with 1:10 Ca/Mg ions ratio, together with high Cu translocation to above-ground plant organs, which suggests its higher potential in phytoremediation.     

Effect of nitrogen concentration,source, and phosphate concentration on accumulation of biomass and calycosin-7-O-β-D-glucoside in Astragalus membranaceus adventitious roots

In Vitro Cellular & Developmental Biology - Plant - Calycosin-7-O-β-D-glucoside (CG), a methoxylated isoflavonoid in Astragalus membranaceus Fisch. (Bunge), has a wide range of biological...     

The influence of Agrobacterium rhizogenes on induction of hairy roots and ß-carboline alkaloids production in Tribulus terrestris L.

We have developed an efficient transformation system for Tribulus terrestris L., an important medicinal plant, using Agrobacterium rhizogenes strains AR15834 and GMI9534 to generate hairy roots. Hairy roots were formed directly from the cut edges of leaf explants 10–14 days after inoculation with the Agrobacterium with highest frequency transformation being 49 %, which was achieved using Agrobacterium rhizogenes AR15834 on hormone-free MS medium after 28 days inoculation. PCR analysis showed that rolB genes of Ri plasmid of A. rhizogenes were integrated and expressed into the genome of transformed hairy roots. Isolated transgenic hairy roots grew rapidly on MS medium supplemented with indole-3-butyric acid. They showed characteristics of transformed roots such as fast growth and high lateral branching in comparison with untransformed roots. Isolated control and transgenic hairy roots grown in liquid medium containing IBA were analyzed to detect ß-carboline alkaloids by High Performance Thin Layer Chromatograghy (HPTLC). Harmine content was estimated to be 1.7 μg g⁻¹ of the dried weight of transgenic hairy root cultures at the end of 50 days of culturing. The transformed roots induced by AR15834 strain, spontaneously, dedifferentiated as callus on MS medium without hormone. Optimum callus induction and shoot regeneration of transformed roots in vitro was achieved on MS medium containing 0.4 mg L⁻¹ naphthaleneacetic acid and 2 mg L⁻¹ 6-benzylaminopurine (BAP) after 50 days. The main objective of this investigation was to establish hairy roots in this plant by using A. rhizogenes to synthesize secondary products at levels comparable to the wild-type roots.     

Alteration of flower color in Iris germanica L. ‘Fire Bride’ through ectopic expression of phytoene synthase gene (crtB) from Pantoea agglomerans

Key message

Genetic modulation of the carotenogenesis in I. germanica ‘Fire Bride’ by ectopic expression of a crtB gene causes several flower parts to develop novel orange and pink colors.

Abstract

Flower color in tall bearded irises (Iris germanica L.) is determined by two distinct biochemical pathways; the carotenoid pathway, which imparts yellow, orange and pink hues and the anthocyanin pathway, which produces blue, violet and maroon flowers. Red-flowered I. germanica do not exist in nature and conventional breeding methods have thus far failed to produce them. With a goal of developing iris cultivars with red flowers, we transformed a pink iris I. germanica, ‘Fire Bride’, with a bacterial phytoene synthase gene (crtB) from Pantoea agglomerans under the control of the promoter region of a gene for capsanthin–capsorubin synthase from Lilium lancifolium (Llccs). This approach aimed to increase the flux of metabolites into the carotenoid biosynthetic pathway and lead to elevated levels of lycopene and darker pink or red flowers. Iris callus tissue ectopically expressing the crtB gene exhibited a color change from yellow to pink-orange and red, due to accumulation of lycopene. Transgenic iris plants, regenerated from the crtB-transgenic calli, showed prominent color changes in the ovaries (green to orange), flower stalk (green to orange), and anthers (white to pink), while the standards and falls showed no significant differences in color when compared to control plants. HPLC and UHPLC analysis confirmed that the color changes were primarily due to the accumulation of lycopene. In this study, we showed that ectopic expression of a crtB can be used to successfully alter the color of certain flower parts in I. germanica ‘Fire Bride’ and produce new flower traits.     

Physical mapping of three fruit ripening genes：Endopolygalacturonase,ACC oxidase and ACC synthase from apple（Malus x domestica） in an apple rootstock A106（Malus sieboldii

The apple rootstock,A106(Malus sieboldii),had 17 bivalents in pollen mother cells at meiotic metaphase 1,and 17 chromosomes in a haploid pollen cell.Karyotypes were prepared from root-tip cells with 2n=34 chromosomes,Seven out of 82 karyotypes(8.5%) showed one pari of satellites at the end of the short arm of chromosome 3.C-bands were shown on 6 pairs of chromosomes 2,4,6,8,14,and 16 near the telomeric regions of short arms.Probes for three ripening-related genes from Malus x domestica:endopolygalacturonase(EPG,0.6kb),ACC oxidase(1.2kb),and ACC synthase(2kb)were hybridized in situ to metaphase chromosomes of A106.Hybridization sites for the EPG gene were observed on the long arm of chromosome 14 in 15 out of 16 replicate spreads and proximal to the centromere of chromosomes 6 and 11.For the ACC oxidase gene,hylridization sites were observed in the telomeric region of the short arm of chromosomes 5 and 11 in 87% and 81% of 16 spreads respectively,proxiaml to the centromere of chromosome 1 in 81% of the spreads,and on the long arm of chromosome 13 in 50% of the spreads. Physical mapping of three fruit ripening genes in an apple rootstock A106.Twenty five spreads were studied for the ACC synthase gene and hybridization sites were observed in the telomeric region of the short arm of chromosome 12 in 96% of the spreads.chromosomes 9 and 10 in 76% of the spreads,and chromosome 17 in 56% of the spreads.     

Silver nanoparticles improved explant disinfection,in vitro growth,runner formation and limited ethylene accumulation during micropropagation of strawberry (Fragaria × ananassa)

Plant Cell, Tissue and Organ Culture (PCTOC) - One of the common problems in strawberry (Fragaria?×?ananassa) micropropagation is the vitrification phenomenon (succulent plantlets,...     

Effect of β-Cyclodextrin Nanoparticle on Biosynthesis of Ocimum basilicum L. Monoterpenes,Sesquiterpenes in a Concentration-Dependent Behavior

Russian Journal of Plant Physiology - Engineered nanoparticles can alter the metabolism pathways and the profiling of metabolites in plants. In this study, we prepared the β-cyclodextrin...     

Recombinant expression,purification, and characterization of polyphenol oxidase 2 (VvPPO2) from “Shine Muscat” (Vitis labruscana Bailey × Vitis vinifera L.)

Polyphenol oxidases (PPOs) catalyze browning reactions in various plant organs, therefore controlling the reactions is important for the food industry. PPOs have been assumed to be involved in skin browning of white grape cultivars; however, the molecular mechanism underlying PPO-mediated browning process remains elusive. We have recently identified a new PPO gene named VvPPO2 from “Shine Muscat” (Vitis labruscana Bailey × V. vinifera L.), and have shown that the gene is transcribed at a higher level than the previously identified VvPPO1 in browning, physiologically disordered berry skins at the maturation stage. In this study, we expressed VvPPO2 in Escherichia coli and, using the purified preparation, revealed unique physicochemical characteristics of the enzyme. Our study opens up a way to not only understand the berry skin browning process but also to elucidate the enzymatic maturation process of grape PPOs.     

Effect of ethylene glycol-bis(β-aminoethyl ether)-N,N-tetraacetic acid (EGTA) on stability and activity of methanogenic granular sludge

Summary The effect of the calcium-specific chelant ethylene glycol-bis(\-aminoethyl ether)-N,N-tetraacetic acid (EGTA) on methanogenic granular sludge from a laboratory-scale upflow anaerobic sludge-blanket (UASB) reactor fed propionate and from a full-scale reactor treating paper-mill waste-water was studied. Upon treatment with EGTA both sludge types showed a decrease in the calcium and phosphorus content and a release of protein and polysaccharides, leading to a decrease in strength of papermill granular sludge and a disintegration of propionate-grown granules. After treatment of propionate-grown granular sludge with high EGTA concentrations, the methanogenic activity with propionate and acetate as test substrates decreased by 88 and 33%, respectively. The marked reduction in propionate oxidation activity may be caused by a disruption of the special juxtapositioning of bacteria in the granules. Offsprint requests to: A. J. B. Zehnder