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Root Tissue (root + tissue)
Selected AbstractsTargeted gene analysis in Ulmus americana and U. pumila tissuesFOREST PATHOLOGY, Issue 2 2008C. Nasmith Summary Steady-state gene expression was compared between Dutch elm disease (DED)-susceptible Ulmus americana and DED-resistant U. pumila callus, leaf midrib, root and inner bark tissues. Stress-related cDNAs including phenylalanine ammonia-lyase (PAL), chitinase (CHT) and polygalacturonase-inhibiting protein (PGIP) were isolated and compared following RT-PCR of elm tissues. Complete CHT and partial PAL and PGIP cDNA transcripts were identified, each displaying sequence variation between elm species. These transcripts were Dig-labelled and subsequently used for northern analyses of the elm tissues. Midrib and root tissue displayed highest steady-state gene expression compared with inner bark and callus tissues. A modified nucleic acid isolation technique was necessary for downstream RNA analyses. Lithium chloride and polyvinylpyrrolidone were critical for efficient removal of polysaccharides and phenolics associated with some of the elm tissues. Steady-state gene expression is discussed in relation to the tissues investigated. The use of tissues other than in vitro callus culture more closely represents the tissues associated with the elm's vascular response to DED. [source] Use of , -Glucuronidase Activity to Quantify the Growth of Fusarium oxysporum f. sp. radicis-lycopersici during Infection of TomatoJOURNAL OF PHYTOPATHOLOGY, Issue 6 2005K. K. Papadopoulou Abstract The , -glucuronidase (gus) reporter gene was integrated into the phytopathogenic fungus Fusarium oxysporum f. sp. radicis-lycopersici (FORL) in a co-transformation experiment using the hygromycin B resistance (hph) gene as selective marker, which resulted in the generation of 10 mitotically stable transformants. One transformant, F30, was selected based on the results of prior detailed characterization of the 10 transformants for growth rate, conidia production and pathogenicity in comparison with the wild-type strain. A strong positive correlation was found between GUS activity and accumulated biomass of in vitro -grown fungus and therefore GUS activity was used to study fungal growth quantitatively in two tomato lines. Although a parallel increase in lesion development and GUS activity was noted for both tomato lines, a correlation between the GUS activity and disease progression was not always possible. Interestingly, the levels of GUS activity obtained for the more resistant line were higher than those obtained for the susceptible line, indicating that disease progression in tomato caused by FORL may not be related only to the amount of fungal biomass within the root tissue. [source] Localized and Systemic Increase of Phenols in Tomato Roots Induced by Glomus versiforme Inhibits Ralstonia solanacearumJOURNAL OF PHYTOPATHOLOGY, Issue 10 2004H. H. Zhu Abstract Ralstonia solanacearum is an important plant pathogen in tropical and subtropical countries. Here, we describe the inhibition of R. solanacearum as a result of increased phenols induced locally or systemically by an arbuscular mycorrhizal (AM) fungus. In pot cultures, R. solanacearum populations in the rhizosphere, on root surfaces and in the xylem were decreased by 26.7, 79.3 and 81.7%, respectively, following inoculation of tomato plants (Lycopersicon esculentum Mill.) with Glomus versiforme Berch. Colonization of the plants by both R. solanacearum and G. versiforme increased the contents of soluble phenols and cell-wall bound phenols in root tissue, but with different patterns. Whereas R. solanacearum preferably promoted the cell-wall bound phenol content, G. versiforme preferably enhanced the soluble phenol content. Split root experiments revealed that R. Solanacearum was inhibited by G. versiforme, and that G. versiforme also increased the phenol content systemically, but to a lesser extent than locally. [source] Reservoir and Non-reservoir Hosts of Bean-Wilt Pathogen, Fusarium oxysporum f. sp. phaseoliJOURNAL OF PHYTOPATHOLOGY, Issue 7-8 2001O. D. Dhingra Abstract The capacity of Fusarium oxysporum f. sp. phaseoli to multiply in the roots of 12 non-host plant species was determined with the objective of selecting potential candidates for crop rotation and/or green manuring in infested bean fields. The plants were inoculated at the seedling stage by a benomyl-resistant mutant of the pathogen using the root-dip technique and transplanted to natural soil. The number of colony forming units/g dry root tissue (CFU/g) was determined at the full bloom stage. Quantitatively, the root colonization differed greatly among the plant species. The roots and lower stem of Dolichos lablab, Phaseolus lunatus, Mucuna aterrima, Canavalia ensiforme and Vigna unguiculata were the most compatible with the pathogen and those of Sorghum bicolor, Crotalaria juncea, Oryza sativa and Zea mays were least compatible. No disease symptoms developed on any plant species. Chlamydospore germination in the rhizosphere also differed significantly among the plant species. There was no correlation between percentage chlamydospore germination in the rhizosphere and extent of root colonization. Most plant species recommended for green manuring in bean fields allowed extensive root and stem colonization by F. o. f. sp. phaseoli and were considered as reservoir hosts. All three of the gramineous species tested and C. juncea were classed as non-reservoir host, because the pathogen did not colonize the stem and its multiplication in the roots was very low. These plant species appear to be good candidates for long-term field evaluation to determine their usefulness in an integrated management of Fusarium bean-wilt. [source] Monitoring the colonization of sugarcane and rice plants by the endophytic diazotrophic bacterium Gluconacetobacter diazotrophicus marked with gfp and gusA reporter genesLETTERS IN APPLIED MICROBIOLOGY, Issue 3 2010L.F.M. Rouws Abstract Aims:, To evaluate the colonization process of sugarcane plantlets and hydroponically grown rice seedlings by Gluconacetobacter diazotrophicus strain PAL5 marked with the gusA and gfp reporter genes. Methods and Results:, Sugarcane plantlets inoculated in vitro with PAL5 carrying the gfp::gusA plasmid pHRGFPGUS did not present green fluorescence, but ,-glucuronidase (GUS)-stained bacteria could be observed inside sugarcane roots. To complement this existing inoculation methodology for micropropagated sugarcane with a more rapid colonization assay, we employed hydroponically grown gnotobiotic rice seedlings to study PAL5,plant interaction. PAL5 could be isolated from the root surface (108 CFU g,1) and from surface-disinfected root and stem tissues (104 CFU g,1) of inoculated plants, suggesting that PAL5 colonized the internal plant tissues. Light microscopy confirmed the presence of bacteria inside the root tissue. After inoculation of rice plantlets with PAL5 marked with the gfp plasmid pHRGFPTC, bright green fluorescent bacteria could be seen colonizing the rice root surface, mainly at the sites of lateral root emergence, at root caps and on root hairs. Conclusion:, The plasmids pHRGFPGUS and pHRGFPTC are valid tools to mark PAL5 and monitor the colonization of micropropagated sugarcane and hydroponic rice seedlings. Significance and Impact of the Study:, These tools are of use to: (i) study PAL5 mutants affected in bacteria,plant interactions, (ii) monitor plant colonization in real time and (iii) distinguish PAL5 from other bacteria during the study of mixed inoculants. [source] High specificity generally characterizes mycorrhizal association in rare lady's slipper orchids, genus CypripediumMOLECULAR ECOLOGY, Issue 2 2005RICHARD P. SHEFFERSON Abstract Lady's slipper orchids (Cypripedium spp.) are rare terrestrial plants that grow throughout the temperate Northern Hemisphere. Like all orchids, they require mycorrhizal fungi for germination and seedling nutrition. The nutritional relationships of adult Cypripedium mycorrhizae are unclear; however, Cypripedium distribution may be limited by mycorrhizal specificity, whether this specificity occurs only during the seedling stage or carries on into adulthood. We attempted to identify the primary mycorrhizal symbionts for 100 Cypripedium plants, and successfully did so with two Cypripedium calceolus, 10 Cypripedium californicum, six Cypripedium candidum, 16 Cypripedium fasciculatum, two Cypripedium guttatum, 12 Cypripedium montanum, and 11 Cypripedium parviflorum plants from a total of 44 populations in Europe and North America, yielding fungal nuclear large subunit and mitochondrial large subunit sequence and RFLP (restriction fragment length polymorphism) data for 59 plants. Because orchid mycorrhizal fungi are typically observed without fruiting structures, we assessed fungal identity through direct PCR (polymerase chain reaction) amplification of fungal genes from mycorrhizally colonized root tissue. Phylogenetic analysis revealed that the great majority of Cypripedium mycorrhizal fungi are members of narrow clades within the fungal family Tulasnellaceae. Rarely occurring root endophytes include members of the Sebacinaceae, Ceratobasidiaceae, and the ascomycetous genus, Phialophora. C. californicum was the only orchid species with apparently low specificity, as it associated with tulasnelloid, ceratobasidioid, and sebacinoid fungi in roughly equal proportion. Our results add support to the growing literature showing that high specificity is not limited to nonphotosynthetic plants, but also occurs in photosynthetic ones. [source] Cloning of nodule-specific cDNAs of Galega orientalisPHYSIOLOGIA PLANTARUM, Issue 4 2002Seppo Kaijalainen Differential display was applied in order to clone cDNAs expressed exclusively or predominantly in nodules, compared to uninoculated root tissue of Galega orientalis. Forty-five fragments were unique for nodule RNA. These fragments were reamplified and cloned. Six of them produced a nodule-specific signal on Northern hybridization. These six fragments were sequenced. Five of the sequenced fragments showed homology to nodulin-gene sequences in databases, among them Vicia faba mRNA for protein showing partial homology with Medicago sativa nodulin-25 (Nms25), Pisum sativum PsN466, V. faba CCP2 and CCP4, P. sativum ENOD3, and Maackia amurensis ENOD2. The remaining sequence had no significant homology with sequences in the databanks. Full-size cDNA for the homologue to V. faba mRNA for the protein showing partial homology with M. sativa nodulin-25 (Nms25) and P. sativum PsN466 were cloned and sequenced. [source] Evaluation of Transgenic Poplars Over-Expressing Enzymes of Glutathione Synthesis for Phytoremediation of CadmiumPLANT BIOLOGY, Issue 6 2002A. Koprivova Abstract: Recently, phytoremediation of soils polluted with heavy metals has received a lot of attention. Since glutathione (GSH) and its derivatives (e.g., phytochelatins) play a major role in plant defence against environmental pollutants, we tested the effects of over-expression of bacterial genes for GSH synthesis in poplar on cadmium accumulation. A pilot experiment with CdCl2 in hydroponics revealed that poplars over-expressing ,-glutamylcysteine synthetase (,-ECS) accumulated significantly more Cd in root tissue than wild type or glutathione synthetase over-expressing poplars. To test the partitioning of Cd in different organs, poplar lines over-expressing ,-ECS in the cytosol and in chloroplasts were treated with 0.2 mM CdCl2 in hydroponics. Significant amounts of Cd were translocated to leaves, but significant differences in Cd accumulation were not observed between transgenic and wild type plants. To evaluate these lines for large-scale phytoremediation of cadmium, plants were treated with 2 mM Cd in soil. Over a four-week period, the poplar plants were able to accumulate up to 5.3 mg Cd. Most remarkably, in young leaves of both transgenic lines, Cd was accumulated to concentrations 2.5 - 3 times higher than in the wild type. The increased allocation of cadmium to the young leaves represents a potentional advantage for the phytoremediation process using the same plants over several vegetation periods. The use of transgenic poplar lines with enhanced glutathione production capacity seems to be of particular advantage in highly polluted soils. [source] Control of Nitrate Uptake by Phloem-Translocated Glutamine in Zea mays L. SeedlingsPLANT BIOLOGY, Issue 4 2002P. Pal'ove-Balang Abstract: The putative role of glutamine, exported from leaves to roots, as a negative feedback signal for nitrate uptake was investigated in Zea mays L. seedlings. Glutamine (Gln) was supplied by immersion of the tip-cut leaves in a concentrated solution. Nitrate (NO3,) uptake was measured by its depletion in amino acid-free medium. The treatment with Gln resulted in a strong inhibition of nitrate uptake rate, accompanied by a significant enrichment of amino compounds in root tissue. The effect of N-availability on NO3, uptake was determined in split-root cultures. The plants were subjected to complete or localized N supply. Inducible NO3, uptake systems were also induced in N-deprived roots when the opposite side of the root system was supplied with KNO3. The inhibitory effect of Gln was unaffected by localized N supply on one side of the split-root. The potential role of Gln in the shoot-to-root control of NO3, uptake is discussed. [source] Preferential expression of a plant cystatin at nematode feeding sites confers resistance to Meloidogyne incognita and Globodera pallidaPLANT BIOTECHNOLOGY JOURNAL, Issue 1 2004Catherine J. Lilley Summary The expression patterns of three promoters preferentially active in the roots of Arabidopsis thaliana have been investigated in transgenic potato plants in response to plant parasitic nematode infection. Promoter regions from the three genes, TUB-1, ARSK1 and RPL16A were linked to the GUS reporter gene and histochemical staining was used to localize expression in potato roots in response to infection with both the potato cyst nematode, Globodera pallida and the root-knot nematode, Meloidogyne incognita. All three promoters directed GUS expression chiefly in root tissue and were strongly up-regulated in the galls induced by feeding M. incognita. Less activity was associated with the syncytial feeding cells of the cyst nematode, although the ARSK1 promoter was highly active in the syncytia of G. pallida infecting soil grown plants. Transgenic potato lines that expressed the cystatin OcI,D86 under the control of the three promoters were evaluated for resistance against Globodera sp. in a field trial and against M. incognita in containment. Resistance to Globodera of 70 ± 4% was achieved with the best line using the ARSK1 promoter with no associated yield penalty. The highest level of partial resistance achieved against M. incognita was 67 ± 9% using the TUB-1 promoter. In both cases this was comparable to the level of resistance achieved using the constitutive cauliflower mosaic virus 35S (CaMV35S) promoter. The results establish the potential for limiting transgene expression in crop plants whilst maintaining efficacy of the nematode defence. [source] Membrane phospholipids as a phosphate reserve: the dynamic nature of phospholipid-to-digalactosyl diacylglycerol exchange in higher plantsPLANT CELL & ENVIRONMENT, Issue 10 2008HENRIK TJELLSTRÖM ABSTRACT It is well established that phosphate deficiency induces the replacement of membrane phospholipid with non-phosphorous lipids in extra-plastidial membranes (e.g. plasma membrane, tonoplast, mitochondria). The predominant replacement lipid is digalactosyl diacylglycerol (DGDG). This paper reports that the phospholipid-to-DGDG replacement is reversible, and that when oat seedlings are re-supplied with radio-labelled phosphate, it is initially recovered primarily in phosphatidylcholine (PC). Within 2 d, the shoot contains more than half of the lipid-associated radiolabel, reflecting phosphate translocation. Oat was also cultivated in different concentrations of phosphate and the DGDG/PC ratio in roots and phospholipase activities in isolated plasma membranes was assayed after different times of cultivation. The DGDG/PC ratio in root tissue correlated more closely with plasma membrane-localized phospholipase D, yielding phosphatidic acid (PA), than with plasma membrane-localized PA phosphatase, the activity that results in a decreased proportion of phospolipids. The lipid degradation data did not reflect a significant involvement of phospholipase C, although a putative phospholipase C analogue, non-specific phospholipase C4 (NPC4), was present in oat roots. The correlation between increased phospholipase D activity and DGDG/PC ratio is consistent with a model where phospholipid-to-DGDG replacement involves formation of PA that readily is removed from the plasma membrane for further degradation elsewhere. [source] Jasmonic acid treatment to part of the root system is consistent with simulated leaf herbivory, diverting recently assimilated carbon towards untreated roots within an hourPLANT CELL & ENVIRONMENT, Issue 9 2008GUNNAR JAKOB HENKES ABSTRACT It is known that shoot application of jasmonic acid (JA) leads to an increased carbon export from leaves to stem and roots, and that root treatment with JA inhibits root growth. Using the radioisotope 11C, we measured JA effects on carbon partitioning in sterile, split-root, barley plants. JA applied to one root half reduced carbon partitioning to the JA-treated tissue within minutes, whereas the untreated side showed a corresponding , but slower , increase. This response was not observed when instead of applying JA, the sink strength of one root half was reduced by cooling it: there was no enhanced partitioning to the untreated roots. The slower response in the JA-untreated roots, and the difference between the effect of JA and temperature, suggest that root JA treatment caused transduction of a signal from the treated roots to the shoot, leading to an increase in carbon allocation from the leaves to the untreated root tissue, as was indeed observed 10 min after the shoot application of JA. This supports the hypothesis that the response of some plant species to both leaf and root herbivores may be the diversion of resources to safer locations. [source] Functional characterization and expression analysis of a glutathione transporter, BjGT1, from Brassica juncea: evidence for regulation by heavy metal exposurePLANT CELL & ENVIRONMENT, Issue 10 2003J. BOGS ABSTRACT Glutathione and its derivatives play an important role in the tolerance of plants against heavy metals. A glutathione transporter, BjGT1 (AJ561120), was cloned and functionally characterized from Brassica juncea, a plant which may be used for phytoremediation. The full-length BjGT1 cDNA showed homology with the high affinity glutathione transporter HGT1 from Saccharomyces cerevisiae and shares 92% identity with a putative glutathione transporter from A. thaliana (At4g16370). When expressed in the S. cerevisiae hgt1, strain, BjGT1 complemented the mutant on medium with glutathione as the only sulphur source and mediated the uptake of [3H]GSH. Immunoblot analysis with a peptide-specific antiserum directed against a C-terminal sequence revealed high BjGT1 expression in leaf tissue and relatively low expression in stem tissue, whereas BjGT1 protein was not detectable in root tissue. The amounts of BjGT1 mRNA and protein were analysed during a 6 d exposure of B. juncea to 25 µm Cd(NO3)2. BjGT1 mRNA was strongly induced by cadmium in stems and leaves. Unexpectedly, the amount of BjGT1 protein in leaves showed a pronounced decrease with a minimum after 96 h of Cd exposure, followed by partial recovery. The strong regulation of BjGT1 by cadmium suggests a role of this glutathione transporter during heavy metal exposure. [source] Soil-borne wheat mosaic virus inclusion bodies: structural, compositional and staining propertiesANNALS OF APPLIED BIOLOGY, Issue 2 2003L J LITTLEFIELD Summary Anatomy and cytochemistry of inclusion bodies induced by Soil-borne wheat mosaic virus infection were studied in roots and leaves to learn more about the nature of inclusions and their roles in pathogenesis. Acid Fuchsin, Giemsa stain, Toluidine Blue and Trypan Blue stains facilitated visualization of inclusion bodies. Combined, simultaneous staining with Acid Fuchsin and Toluidine Blue clearly differentiated inclusion bodies from host nuclei. The overall anatomy, composition and structure of virus inclusions in leaves and roots were generally similar, as shown by phase contrast, differential interference contrast, epifluorescence, laser scanning confocal and transmission electron microscopy. Both were often closely associated with host nuclei; both were comprised of intertwined masses of tubular material, presumably endoplasmic reticulum, and in which varied numbers and sizes of vacuolar cavities occurred. Leaf inclusions, however, were typically larger and more vacuolate than those in roots. Lipids were found to be significant constituents of both the tubular and vacuolar components of inclusions, indicated by positive staining with Nile Red and Sudan Black. Inclusion bodies in both leaves and roots lost their structural and compositional integrity, eventually becoming disorganized and devoid of clearly identifiable components as host tissue aged and symptom expression advanced. Significant results of this study include the first published examination of virus inclusion bodies in root tissue, the degree of structural detail of inclusion body anatomy revealed by laser scanning confocal microscopy and the presence of an extensive lipid component in virus inclusion bodies. [source] Alkaloid production in Vernonia cinerea: Callus, cell suspension and root culturesBIOTECHNOLOGY JOURNAL, Issue 8 2007Priti Maheshwari Abstract Fast-growing callus, cell suspension and root cultures of Vernonia cinerea, a medicinal plant, were analyzed for the presence of alkaloids. Callus and root cultures were established from young leaf explants in Murashige and Skoog (MS) basal media supplemented with combinations of auxins and cytokinins, whereas cell suspension cultures were established from callus cultures. Maximum biomass of callus, cell suspension and root cultures were obtained in the medium supplemented with 1 mg/L ,-naphthaleneacetic acid (NAA) and 5 mg/L benzylaminopurine (BA), 1.0 mg/L NAA and 0.1 mg/L BA and 1.5 mg/L NAA, respectively. The 5-week-old callus cultures resulted in maximum biomass and alkaloid contents (750 ,g/g). Cell suspension growth and alkaloid contents were maximal in 20-day-old cultures and alkaloid contents were 1.15 mg/g. A 0.2-g sample of root tissue regenerated in semi-solid medium upon transfer to liquid MS medium containing 1.5 mg/L NAA regenerated a maximum increase in biomass of 6.3-fold over a period of 5 weeks. The highest root growth and alkaloid contents of 2 mg/g dry weight were obtained in 5-week-old cultures. Maximum alkaloid contents were obtained in root cultures in vitro compared to all others including the alkaloid content of in vivo obtained with aerial parts and roots (800 ,g/g and 1.2 mg/g dry weight, respectively) of V. cinerea. [source] Endochitinase activity in the apoplastic fluid of Phellinus weirii -infected Douglas-fir and its association with over wintering and antifreeze activityFOREST PATHOLOGY, Issue 5 2003A. Zamani Summary Extracellular proteins were extracted from Phellinus weirii infected Douglas-fir (Pseudotsuga menziesii var. menziesii) roots and needles to examine endochitinase activity. Chitinases have been associated with the plant's defence response against fungal attack because they hydrolyse chitin, a structural component of fungal cell walls. Protein separation using sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE) followed by Western immunoblot analysis using a polyclonal antibody specific to an endochitinase-like protein (ECP) resulted in the detection of up to three polypeptides between 27 and 30 kDa in size. Two-dimensional gel electrophoresis (2-D PAGE) followed by Western immunoblot analysis revealed that the apoplastic fluid contained multiple ECP isoforms with isoelectric points (pIs) ranging from 5.3 to 5.8 and molecular masses of 27,30 kDa. Chitinase activity in needle and root tissues was measured spectrophotometrically using a colorimetric assay. A gel overlay technique using glycol chitin as a substrate for endochitinase was applied to confirm that the ECP antibody detected an enzymatically active protein. The apoplastic fluid collected from P. weirii -infected winter Douglas-fir needles showed anti-freeze activity and seasonal analysis of needle tissue showed some evidence of ECP accumulation in winter months. ECP was distributed systemically throughout the tree. Increased levels of endochitinase activity in the region of P. weirii infection supports a physiological role for ECP in the plant defence response. Résumé Les protéines extra-cellulaires ont été extraites des racines et aiguilles de douglas (Pseudotsuga menziesii var menziesii) infectés par Phellinus weirii (Murr.) Gilbn., pour étudier l'activité endochitinase. Les chitinases ont été associées aux réactions de défense des plantes contre les attaques fongiques parce-qu'elles hydrolysent la chitine, un composant de la paroi des cellules fongiques. La séparation des protéines, réalisée par électrophorèse en gel de polyacrylamide avec sodium dodecyl sulfate (SDS-PAGE), suivie par une analyse par Western immunoblot en utilisant un anticorps polyclonal spécifique d'une protéine de type endochitinase (ECP), a permis la détection de 3 polypeptides de taille comprise entre 27 et 30 kDa. Une électrophorèse sur gel en 2-dimensions (2-D PAGE) suivie par une analyse par Western immunoblot a révélé que le fluide apoplastique contient de multiples isoformes d'ECP avec des pI dans une gamme de 5.3 à 5.8 et des masses moléculaires de 27 à 30 kDa. L'activité chitinase dans les aiguilles et tissus racinaires a été mesurée par spectrophotométrie par une méthode colorimétrique. Une technique d'overlay utilisant de la chitine glycol comme substrat de l'endochitinase a été appliquée pour confirmer que l'anticorps ECP avait détecté une protéine active du point de vue enzymatique. Le fluide apoplastique d'aiguilles récoltées en hiver sur des douglas infectés par P. weirii a montré une activité antigel et l'analyse saisonnière des tissus foliaires a montré une certaine accumulation d'ECP pendant l'hiver. L'ECP est répartie de façon systémique dans l'ensemble de l'arbre. Les niveaux accrus d'activité endochitinase dans la zone infectée par P. weirii suggère un rôle physiologique de l'ECP dans les réactions de défense de la plante. Zusammenfassung Aus Wurzeln und Nadeln von mit Phellinus weirii infizierten Douglasien (Pseudotsuga menziesii var. menziesii) wurden extrazelluläre Proteine extrahiert, um die Endochitinase-Aktivität zu bestimmen. Chitinasen werden mit der pflanzlichen Abwehrreaktion auf Pilzinfektionen in Verbindung gebracht, da sie Chitin, eine Strukturkomponente der pilzlichen Zellwand, hydrolysieren. Die Proteine wurden mit Natrium-Dodecyl-Sulfat-Polyacrylamid-Gelelektrophorese (SDS-PAGE) getrennt, gefolgt von einer Western Immunoblot-Analyse mit einem gegen ein Endochitinase-ähnliches Protein (ECP) spezifischen polyklonalen Antikörper. Hiermit liessen sich bis zu drei Polypeptide zwischen 27-30 kDa nachweisen. Eine zweidimensionale Gelelektrophorese (2-D PAGE) mit anschliessender Western Immunoblot-Analyse ergab, dass die Apoplastenflüssigkeit multiple ECP-Isoformen enthielt (mit pIs von 5,3 bis 5,8 und Molekularmassen von 27 bis 30 kDa). Die Chitinase-Aktivität wurde auch im Nadel- und Wurzelgewebe spektrophotometrisch mit einer Farbreaktion gemessen. Um sicher zu stellen, dass der ECP-Antikörper ein enzymatisch aktives Protein nachwies, wurde eine Gel-Overlay-Methode verwendet, mit Glycolchitin als Substrat für die Endochitinase. Die Apoplastenflüssigkeit der Nadeln von mit P. weirii infizierten Douglasien zeigte in Winterzustand eine Antifrost-Aktivität, ihre Analyse während des gesamten Jahres ergab aber keine Hinweise auf eine ECP-Anreicherung während der Wintermonate. ECP war systemisch im gesamten Baum enthalten. Die erhöhte Endochitinase-Aktivität in Bereichen mit P. weirii -Infektion lässt auf eine physiologische Rolle von ECP in der Pflanzenabwehr schliessen. [source] Phenolic Acid Content and Composition in Leaves and Roots of Common Commercial Sweetpotato (Ipomea batatas L.) Cultivars in the United StatesJOURNAL OF FOOD SCIENCE, Issue 6 2007V.-D. Truong ABSTRACT:, Phenolic acids in commercially important sweet potato cultivars grown in the United States were analyzed using reversed-phase high-performance liquid chromatography (HPLC). Caffeic acid, chlorogenic acid, 4,5-di-O-caffeoylquinic acid, 3,5-di-O-caffeoylquinic acid, and 3,4-di-O-caffeoylquinic acid were well separated with an isocratic elution in less than 25 min compared to about 120 min for analyzing and re-equilibrating the column with a gradient method. The isocratic elution order of these caffeoylquinic acid derivatives was confirmed by LC-MS/MS. Chlorogenic acid was the highest in root tissues, while 3,5-di-O-caffeoylquinic acid and/or 4,5-di-O-caffeoylquinic acid were predominant in the leaves. Steam cooking resulted in statistically nonsignificant increases in the concentration of total phenolics and all the individual phenolic acids identified. Sweetpotato leaves had the highest phenolic acid content followed by the peel, whole root, and flesh tissues. However, there was no significant difference in the total phenolic content and antioxidant activity between purees made from the whole and peeled sweet potatoes. [source] Identity and Pathogenicity of Fungi Associated with Root and Crown Rot of Soft Red Winter Wheat Grown on the Upper Coastal Plain Land Resource Area of MississippiJOURNAL OF PHYTOPATHOLOGY, Issue 2 2000M. S. Gonzalez Seedling stand, disease severity and fungal incidence were determined from untreated ,Wakefield' soft red winter wheat planted on a Leeper silty clay loam in field tests conducted at the Mississippi Agricultural and Forestry Experiment Station, Plant Science Research Center, Mississippi State University, Starkville, Mississippi during the 1996,97 and 1997,98 growing seasons. Seedling stand was reduced by 40% each year in plots established with untreated seed. Cochliobolus sativus was the most frequently isolated fungus. Fusarium acuminatum, Fusarium equiseti and Fusarium solani were the most prevalent Fusarium spp. Seven other Fusarium spp. and 23 species of other fungal genera were isolated. Pathogenicity tests with three isolates each of C. sativus, Cochliobolus spicifer, F. acuminatum, F. solani, F. equiseti, Fusarium compactum, Embellisia chlamydospora and Microdochium bolleyi were performed in test tube culture and two isolates each of C. sativus, C. spicifer, F. acuminatum, E. chlamydospora and M. bolleyi under greenhouse conditions. In test tubes and in the greenhouse, seedlings infected with isolates of C. sativus developed seedling blight, discoloration and necrosis, primarily in seminal roots and crowns. In the greenhouse, C. sativus induced lesions on the lower leaf sheath and reduced seedling height, seedling emergence, dry and fresh weight of roots and shoots. Isolates of F. acuminatum, F. solani, F. equiseti, F. compactum, E. chlamydospora and M. bolleyi induced slight to moderate orange to light-brown discoloration of crown and seminal roots in test tubes. Cochliobolus spicifer isolates had the most pre-emergence activity, inducing black root discoloration and root pruning of wheat seedlings and reducing seedling emergence, root fresh weight and shoot dry weight. In the greenhouse, F. acuminatum reduced seedling height, seedling emergence and root and shoot dry weights. Microdochium bolleyi and E. chlamydospora reduced fresh and dry weight of roots, plant emergence and shoot dry weight. Fusarium acuminatum and C. spicifer reduced the growth rate of wheat seedlings. All fungi evaluated showed increased disease severity compared to the untreated control. The high frequency of isolation of C. sativus from crown and root tissues can be partially explained by the dry, warm conditions during the early stages of wheat seedling development in the Upper Coastal Plain Land Resource Area of Mississippi. Zusammenfassung Die Auflaufrate von Sämlingen, die Stärke des Krank-heitsbefalls sowie die Häufigkeit von Pilzarten wurden bei nicht behandelten roten Weichwinterweizen der Sorte Wakefield ermittelt, welche in einem Leeper schlammigen Tonboden an der Mississippi Agricultural & Forestry Experiment Station, Plant Science Research Center, Mississippi State University, Starkville, Mississippi in der 1996,97 und 1997,98 Saison gesät worden waren. In beiden Jahren wurde die Auflaufrate von nicht behandeltem Saatgut um 40% reduziert. Cochliobolus sativus wurde am häufigsten isoliert. Fusarium acuminatum, Fusarium equiseti und Fusarium solani waren die überwiegenden Fusarium spp. Außierdem wurden sieben weitere Fusarium spp. sowie 23 weitere Pilzarten isoliert. Pathogenitätstests mit je 3 Isolaten von C. sativus, Cochliobolus spicifer, F. acuminatum, F. solani, F. equiseti, Fusarium compactum, Embellisia chlamydospora und Microdochiumbolleyi wurden in Reagenzröhrchen durchgeführt, sowie mit je 2 Isolaten von C. sativus, C. spicifer, F. acuminatum, E. chlamydospora und M. bolleyi unter Gewächshausbedingungen. Sowohl in den Reagenzröhrchen als auch im Gewächshaus entwickelten Sämlinge, die mit C. sativus inokuliert worden waren, eine Fäule, Verfärbung sowie Nekrosis, hauptsächlich in den sekundären Wurzeln und in den Halmbasen. Unter Gewächshausbedingungen verursachte C. sativus außierdem Läsionen der unteren Blattscheide sowie eine Reduzierung des Sämlingswachstums, des Sämlingsauflaufs, des Trocken-und Frischgewichts der Wurzeln und Sprossen. Im Reagenzröhrchentest induzierten Isolate von F. acuminatum, F. solani, F. equiseti, F. compactum, E. chlamydospora und M. bolleyieine schwache bis mäßiige orange bis hell braune Verfärbung des Halmbasis und der Sekundärwurzeln. Isolate von C. spicifer besaßien die höchste Vorauflaufaktivität und induzierten eine Verschwärzung und Verkürzung der Wurzeln sowie eine Reduzierung des Sämlingsauflaufs, des Wurzelfrischgewichts sowie des Sproitrockengewichts. Unter Gewächshausbedingungen reduzierte F. acuminatum die Sämlingshöhe, die Auflaufrate sowie das Trockengewicht der Wurzeln und Sproien. Microdochium bolleyi und E. chlamydospora reduzierten das Frisch-und Trockengewicht der Wurzeln, die Auflaufrate sowie das Sproßitrockengewicht. Die Wachstumsrate der Sämlinge wurde durch F. acuminatum und C. spicifer reduziert. Alle untersuchten Pilzarten erhöhten die Befallsstärke verglichen mit der unbehandelten Kontrolle. Die hohe Isolierungsrate von C. sativus aus dem Halmbasis-und Wurzelgewebe kann zum Teil dadurch erklärt werden, dass während der Frühentwicklungsphase der Sämlinge trockene und warme Wachstumsbedingungen in diesem Gebiet herrschten. [source] Melatonin in Glycyrrhiza uralensis: response of plant roots to spectral quality of light and UV-B radiationJOURNAL OF PINEAL RESEARCH, Issue 2 2006F. Afreen Abstract:, Melatonin (N-acetyl-5-methoxytryptamine) is known to be synthesized and secreted by the pineal gland in vertebrates. Evidence for the occurrence of melatonin in the roots of Glycyrrhiza uralensis plants and the response of this plant to the spectral quality of light including red, blue and white light (control) and UV-B radiation (280,315 nm) for the synthesis of melatonin were investigated. Melatonin was extracted and quantified in seed, root, leaf and stem tissues and results revealed that the root tissues contained the highest concentration of melatonin; melatonin concentrations also increased with plant development. After 3 months of growth under red, blue and white fluorescent lamps, the melatonin concentrations were highest in red light exposed plants and varied depending on the wavelength of light spectrum in the following order red , blue , white light. Interestingly, in a more mature plant (6 months) melatonin concentration was increased considerably; the increments in concentration were X4, X5 and X3 in 6-month-old red, blue and white light exposed (control) plants, respectively. The difference in melatonin concentrations between blue and white light exposed (control) plants was not significant. The concentration of melatonin quantified in the root tissues was highest in the plants exposed to high intensity UV-B radiation for 3 days followed by low intensity UV-B radiation for 15 days. The reduction of melatonin under longer periods of UV-B exposure indicates that melatonin synthesis may be related to the integrated (intensity and duration) value of UV-B irradiation. Melatonin in G. uralensis plant is presumably for protection against oxidative damage caused as a response to UV irradiation. [source] Colonization process of olive tissues by Verticillium dahliae and its in planta interaction with the biocontrol root endophyte Pseudomonas fluorescens PICF7MICROBIAL BIOTECHNOLOGY, Issue 4 2009Pilar Prieto Summary The colonization process of Olea europaea by the defoliating pathotype of Verticillium dahliae, and the in planta interaction with the endophytic, biocontrol strain Pseudomonas fluorescens PICF7 were determined. Differential fluorescent protein tagging was used for the simultaneous visualization of P. fluorescens PICF7 and V. dahliae in olive tissues. Olive plants were bacterized with PICF7 and then transferred to V. dahliae -infested soil. Monitoring olive colonization events by V. dahliae and its interaction with PICF7 was conducted using a non-gnotobiotic system, confocal laser scanner microscopy and tissue vibratoming sections. A yellow fluorescently tagged V. dahliae derivative (VDAT-36I) was obtained by Agrobacterium tumefaciens -mediated transformation. Isolate VDAT-36I quickly colonized olive root surface, successfully invaded root cortex and vascular tissues via macro- and micro-breakages, and progressed to the aerial parts of the plant through xylem vessel cells. Strain PICF7 used root hairs as preferred penetration site, and once established on/in root tissues, hindered pathogen colonization. For the first time using this approach, the entire colonization process of a woody plant by V. dahliae is reported. Early and localized root surface and root endophytic colonization by P. fluorescens PICF7 is needed to impair full progress of verticillium wilt epidemics in olive. [source] Frozen in time: a new method using cryo-scanning electron microscopy to visualize root,fungal interactionsNEW PHYTOLOGIST, Issue 2 2006Steve Refshauge Summary ,,A new method of sample preparation for cryo-scanning electron microscopy was used to visualize internal infection of wheat (Triticum aestivum) roots by the pathogenic fungus Rhizoctonia solani AG-8. The new method retained fungal hyphae and root cells in situ in disintegrating root tissues, thus avoiding the distortions that can be introduced by conventional preparation by chemical fixation, dehydration and embedding. ,,Infected roots frozen in liquid nitrogen were cryo-planed and etched (sublimed) at ,80°C for a critical length of time (up to 9 min) in the microscope column to reveal plant and fungal structures in three dimensions. ,,Root and fungal structures were well preserved irrespective of infection severity. Root and hyphal cell walls were clearly seen and hyphal architecture within and between root cells was preserved. ,,This rapid method permits three-dimensional in situ visualization of fungal invasion within roots and has broad application for examination of diseases caused by other necrotrophic fungi. [source] Thermoperiod affects the diurnal cycle of nitrate reductase expression and activity in pineapple plants by modulating the endogenous levels of cytokininsPHYSIOLOGIA PLANTARUM, Issue 3 2009Luciano Freschi Nitrate reductase (NR, EC 1.6.6.1) activity in higher plants is regulated by a variety of environmental factors and oscillates with a characteristic diurnal rhythm. In this study, we have demonstrated that the diurnal cycle of NR expression and activity in pineapple (Ananas comosus, cv. Smooth Cayenne) can be strongly modified by changes in the day/night temperature regime. Plants grown under constant temperature (28°C light/dark) showed a marked increase in the shoot NR activity (NRA) during the first half of the light period, whereas under thermoperiodic conditions (28°C light/15°C dark) significant elevations in the NRA were detected only in the root tissues at night. Under both conditions, increases in NR transcript levels occurred synchronically about 4 h prior to the corresponding elevation of the NRA. Diurnal analysis of endogenous cytokinins indicated that transitory increases in the levels of zeatin, zeatin riboside and isopentenyladenine riboside coincided with the accumulation of NR transcripts and preceded the rise of NRA in the shoot during the day and in the root at night, suggesting these hormones as mediators of the temperature-induced modifications of the NR cycle. Moreover, these cytokinins also induced NRA in pineapple when applied exogenously. Altogether, these results provide evidence that thermoperiodism can modify the diurnal cycle of NR expression and activity in pineapple both temporally and spatially, possibly by modulating the day/night changes in the cytokinin levels. A potential relationship between the day/night NR cycle and the photosynthetic pathway performed by the pineapple plants (C3 or CAM) is also discussed. [source] Osmoprotectant , -alanine betaine synthesis in the Plumbaginaceae: S -adenosyl- l -methionine dependent N -methylation of , -alanine to its betaine is via N -methyl and N,N -dimethyl , -alaninesPHYSIOLOGIA PLANTARUM, Issue 3 2000Bala Rathinasabapathi , -Alanine betaine is an osmoprotective compound accumulated by most members of the plant family Plumbaginaceae. Leaf and root tissues of Limonium latifolium known to accumulate , -alanine betaine readily convert supplied , -alanine to , -alanine betaine. To identify the intermediates and the enzymes involved in , -alanine betaine synthesis, radiotracer experiments using [ C] formate were employed. These studies demonstrate that , -alanine betaine is synthesized from , -alanine via N -methyl and N,N- dimethyl , -alanines. A rapid and sensitive radiometric assay was developed to measure N -methyltransferase (NMT) activities by using [methyl- 14C] or [methyl- 3H] S -adenosyl- l -methionine (AdoMet) as the methyl donor. Leaf extracts from , -alanine betaine accumulators ,Armeria maritima, L. latifolium and L. ramosissimum, had detectable NMT activities while none were found in L. perezii, a species that does not accumulate , -alanine betaine. The NMT activities were further characterized from the leaves of L. latifolium. The activities had a pH optimum of 8.0, were soluble and inhibited by S -adenosyl- l -homocysteine. Extractable activities were similar from plants grown under control and salinity stress conditions. Radiolabeling with [ C] l -aspartic acid indicated that, unlike in bacteria, decarboxylation of l -aspartic acid is not the source of , -alanine in the Plumbaginaceae. [source] Heterologous expression of Arabidopsis H+ -pyrophosphatase enhances salt tolerance in transgenic creeping bentgrass (Agrostis stolonifera L.)PLANT CELL & ENVIRONMENT, Issue 2 2010ZHIGANG LI ABSTRACT The Arabidopsis vacuolar H+ -pyrophosphatase (AVP1), when over-expressed in transgenic (TG) plants, regulates root and shoot development via facilitation of auxin flux, and enhances plant resistance to salt and drought stresses. Here, we report that TG perennial creeping bentgrass plants over-expressing AVP1 exhibited improved resistance to salinity than wild-type (WT) controls. Compared to WT plants, TGs grew well in the presence of 100 mm NaCl, and exhibited higher tolerance and faster recovery from damages from exposure to 200 and 300 mm NaCl. The improved performance of the TG plants was associated with higher relative water content (RWC), higher Na+ uptake and lower solute leakage in leaf tissues, and with higher concentrations of Na+, K+, Cl - and total phosphorus in root tissues. Under salt stress, proline content was increased in both WT and TG plants, but more significantly in TGs. Moreover, TG plants exhibited greater biomass production than WT controls under both normal and elevated salinity conditions. When subjected to salt stress, fresh (FW) and dry weights (DW) of both leaves and roots decreased more significantly in WT than in TG plants. Our results demonstrated the great potential of genetic manipulation of vacuolar H+ -pyrophosphatase expression in TG perennial species for improvement of plant abiotic stress resistance. [source] Sequencing over 13 000 expressed sequence tags from six subtractive cDNA libraries of wild and modern wheats following slow drought stressPLANT CELL & ENVIRONMENT, Issue 3 2009NESLIHAN Z. ERGEN ABSTRACT A deeper understanding of the drought response and genetic improvement of the cultivated crops for better tolerance requires attention because of the complexity of the drought response syndrome and the loss of genetic diversity during domestication. We initially screened about 200 wild emmer wheat genotypes and then focused on 26 of these lines, which led to the selection of two genotypes with contrasting responses to water deficiency. Six subtractive cDNA libraries were constructed, and over 13 000 expressed sequence tags (ESTs) were sequenced using leaf and root tissues of wild emmer wheat genotypes TR39477 (tolerant) and TTD-22 (sensitive), and modern wheat variety Kiziltan drought stressed for 7 d. Clustering and assembly of ESTs resulted in 2376 unique sequences (1159 without hypothetical proteins and no hits), 75% of which were represented only once. At this level of EST sampling, each tissue shared a very low percentage of transcripts (13,26%). The data obtained indicated that the genotypes shared common elements of drought stress as well as distinctly differential expression patterns that might be illustrative of their contrasting ability to tolerate water deficiencies. The new EST data generated here provide a highly diverse and rich source for gene discovery in wheat and other grasses. [source] Cyclitols and carbohydrates in leaves and roots of 13 Eucalyptus species suggest contrasting physiological responses to water deficitPLANT CELL & ENVIRONMENT, Issue 11 2006ANDREW MERCHANT ABSTRACT In many tree species, physiological adaptations to drought include the accumulation of osmotically active substances and/or the presence of particular compatible solutes, among them cyclitols. Recently, the cyclitol quercitol was identified in species of Eucalyptus, a diverse genus whose speciation is probably driven by adaptation to water availability. We subjected seedlings of 13 Eucalyptus species from different ecosystems (,mesic' and ,xeric') and different sub-generic taxonomic groups to 10 weeks of water deficit (WD) treatment. Pre-dawn water potentials (,pdwn) and relative water content (RWC) were determined in shoots, and total osmolality, soluble low-molecular-weight carbohydrates and cyclitols were measured in leaves and roots. Responses to water deficit followed two distinct patterns: Eucalyptus species from ,mesic' environments adjusted concentrations of sucrose (through increased levels of sucrose and decreases in RWC) in response to water deficit, whereas ,xeric' species increased concentrations of quercitol (through reductions in RWC). In root tissues, only species from xeric environments contained high levels of quercitol and mannitol, increasing under WD conditions. We suggest that the former (mesic) strategy may be beneficial to respond to short-lasting drought conditions, because sucrose is easily metabolized, whereas the latter (xeric) strategy may relate to an effective acclimation to longer-lasting drought. These physiological response groups are also related to taxonomic groups within the genus. [source] Induction of root-mat symptoms on cucumber plants by Rhizobium, but not by Ochrobactrum or Sinorhizobium, harbouring a cucumopine Ri plasmidPLANT PATHOLOGY, Issue 6 2005S. A. Weller Ochrobactrum CSL 2573, Rhizobium CSL 2411 and Sinorhizobium CSL 2611 strains harbouring the Agrobacterium cucumopine Ri plasmid (pRi), previously were shown to induce root-mat symptoms in an in vitro cucumber cotyledon assay. In whole-plant, rockwool-grown cucumber host tests Rhizobium CSL 2411 was shown to be as efficient an inducer of root-mat symptoms as the virulent Agrobacterium radiobacter strain NCPPB 4042, which also harbours a cucumopine pRi. Conjugal transfer of pRi to ingressing, avirulent Agrobacterium isolates was observed within root tissues with symptoms. Ochrobactrum CSL 2573 and Sinorhizobium CSL 2611 were not able to induce root-mat symptoms on plants. Rhizobium CSL 2411 and Ochrobactrum CSL 2573 were reisolated from inoculated plants, but Sinorhizobium CSL 2611 was not detected or isolated from inoculated plants 68 days after inoculation. It was postulated that the differences in pathogenicity observed between the in vitro and in situ host tests were caused by a lack of proper attachment to inoculated root tissues by pRi-harbouring Ochrobactrum and Sinorhizobium in the whole-plant host tests. [source] SVISS , a novel transient gene silencing system for gene function discovery and validation in tobacco plantsTHE PLANT JOURNAL, Issue 5 2002Véronique Gosselé Summary We developed a novel, two-component transient gene silencing system in which the satellite tobacco mosaic virus (STMV) is used as vector for the delivery of inhibitory RNA into tobacco plants and the tobacco mosaic virus strain U2 (TMV-U2) is used as helper virus for supplying replication and movement proteins in trans. The main advantage of the system is that by uncoupling virus replication components from silencing induction components, the intensity of silencing becomes more pronounced. We call this system satellite virus-induced silencing system (SVISS) and will demonstrate here its robustness, speed and effectiveness. We were able to obtain pronounced and severe knockout phenotypes for a range of targeted endogenous genes belonging to various biochemical pathways and expressed in different plant tissues, such as genes involved in leaf and flower pigmentation, genes for cell wall synthesis in leaf, stem and root tissues or a ubiquitous RNA polymerase gene. By tandem insertion of more than one target gene sequence into the vector, we were able to induce simultaneous knockouts of an endogenous gene and a transgene. SVISS is the first transient gene silencing system for Nicotiana tabacum, which is a genetically well-characterized bridging species for the Solanaceae plant family. [source] Effect of Elicitation on Growth, Respiration, and Nutrient Uptake of Root and Cell Suspension Cultures of HyoscyamusmuticusBIOTECHNOLOGY PROGRESS, Issue 2 2002Edgard B. Carvalho The elicitation of Hyoscyamus muticus root and cell suspension cultures by fungal elicitor from Rhizoctonia solani causes dramatic changes in respiration, nutrient yields, and growth. Cells and mature root tissues have similar specific oxygen uptake rates (SOUR) before and after the onset of the elicitation process. Cell suspension SOUR were 11 and 18 ,mol O2/g FW·h for non-elicited control and elicited cultures, respectively. Mature root SOUR were 11 and 24 ,mol O2/g FW·h for control and elicited tissue, respectively. Tissue growth is significantly reduced upon the addition of elicitor to these cultures. Inorganic yield remains fairly constant, whereas yield on sugar is reduced from 0.532 to 0.352 g dry biomass per g sugar for roots and 0.614 to 0.440 g dry biomass per g sugar for cells. This reduction in yield results from increased energy requirements for the defense response. Growth reduction is reflected in a reduction in root meristem (tip) SOUR, which decreased from 189 to 70 ,mol O2/g FW·h upon elicitation. Therefore, despite the increase in total respiration, the maximum local oxygen fluxes are reduced as a result of the reduction in metabolic activity at the meristem. This distribution of oxygen uptake throughout the mature tissue could reduce mass transfer requirements during elicited production. However, this was not found to be the case for sesquiterpene elicitation, where production of lubimin and solavetivone were found to increase linearly up to oxygen partial pressures of 40% O2 in air. SOUR is shown to similarly increase in both bubble column and tubular reactors despite severe mass transfer limitations, suggesting the possibility of metabolically induced increases in tissue convective transport during elicitation. [source] | ||||||||||||||||||||||||||||