Showing papers in "Biologia Plantarum in 2012"

The identification of flavonoids and the expression of genes of anthocyanin biosynthesis in the chrysanthemum flowers

Abstract: In order to provide additional information on the coloration of chrysanthemum flowers, the flavonoid composition and the expression of six structural genes involved in anthocyanin pathway in the ray florets of a pink flowering (cv. H5) and two white flowering (cvs. Keikai and Jinba) Chrysanthemum grandiflorum cultivars were examined. HPLCDAD/ESI-MSn analysis showed that cyanidin 3-O-(6″-O-malonylglucoside) and cyanidin 3-O-(3″,6″-O-dimalonylglucoside) were the two major flavonoids presented in H5, while white flowering cultivars contained flavones instead of anthocyanins. Nine flavone derivatives were detected in the three cultivars, the amount of each flavone varied upon cultivars, and seven of these were identified as luteolin 7-O-arabinosylglucuronide, apigenin 7-O-glucoside, luteolin 7-O-malonylglucoside, apigenin 7-O-malonylglucoside, chrysoeriol 7-O-malonylglucoside, acacetin 7-O-rutinoside and acacetin 7-O-malonylglucoside. The two white flowering cultivars showed similar total flavonoid content, which was about two fold higher than that in H5. A high expression of the genes encoding dihydroflavonol 4-reductase and 3-O-glucosyltransferase was detected only in H5 but not in Keikai or Jinba. Chalcone synthase, chalcone isomerase, flavanone 3-hydroxylase, and flavonoid 3′-hydroxylase were expressed in all flowers, suggesting that the lack of anthocyanin in white flowering cultivars cannot be due to any blockage of their expression.

Aluminum-induced changes in reactive oxygen species accumulation, lipid peroxidation and antioxidant capacity in wheat root tips

Abstract: The present study investigated the effects of aluminum on lipid peroxidation, accumulation of reactive oxygen species and antioxidative defense systems in root tips of wheat (Triticum aestivum L) seedlings Exposure to 30 μM Al increased contents of malondialdehyde, H2O2, suproxide radical and Evans blue uptake in both genotypes, with increases being greater in Al-sensitive genotype Yangmai-5 than in Al-tolerant genotype Jian-864 In addition, Al treatment increased the activity of superoxide dismutase (SOD), peroxidase (POD), catalase (CAT), ascorbate peroxidase (APX), monodehydroascorbate reductase (MDHAR), glutathione reductase (GR) and glutathione peroxidase (GPX), as well as the contents of ascorbate (AsA) and glutathione (GSH) in both genotypes The increased activities SOD and POD were greater in Yangmai-5 than in Jian-864, whereas the opposite was true for the activities of CAT, APX, MDHAR, GR and GPX and the contents of AsA and GSH Consequently, the antioxidant capacity in terms of 2,2-diphenyl-1-picrylhydrazyl (DPPH)-radical scavenging activity and ferric reducing/antioxidant power (FRAP) was greater in Jian-864 than in Yangmai-5

Amelioration of salt-induced oxidative stress in eggplant by application of 24-epibrassinolide

Abstract: The effects of exogenous 24-epibrassinolide (EBR) on the growth, oxidative damage, antioxidant system and ion contents in eggplant (Solanum melongena L.) seedlings under salt stress were investigated. Eggplant seedlings were exposed to 90 mM NaCl with 0, 0.025, 0.05, 0.10 and 0.20 mg dm−3 EBR for 10 d. EBR, especially at concentration 0.05 mg dm−3, alleviated growth suppression caused by NaCl stress, decreased electrolyte leakage, superoxide production and content of malondialdehyde and H2O2 in NaCl-treated plants. EBR also increased activities of superoxide dismutase, guaiacol peroxidase, catalase and ascorbate peroxidase and the contents of ascorbic acid and reduced glutathione. Furthermore, we also found that Na+, Cl− contents were decreased, K+, Ca2+ contents and K+/Na+, Ca2+/Na+ ratios were increased in the presence of EBR under salt stress.

Pretreatment with 5-aminolevulinic acid mitigates heat stress of cucumber leaves

Abstract: Cucumber seedlings were pretreated with 3 μM 5-aminolevulinic acid (ALA) followed by cultivation at normal (25/18 °C) or high (42/38 °C) day/night temperature to investigate the protective effects of ALA on heat stress in plants. Heat elevated the contents of malondiadehyde (MDA), superoxide radical (O2.−) and hydrogen peroxide (H2O2) in leaves of all plants but less in ALA-pretreated plants. Heat treatment resulted in higher antioxidant enzyme activities and proline and soluble sugar contents and weaker growth inhibition in ALA-pretreated plants than in those treated with heat alone. These results indicate that ALA pretreatment increased the tolerance of seedlings to heat stress.

Ascorbate deficient semi-dwarf asfL1 mutant of Lathyrus sativus exhibits alterations in antioxidant defense

Abstract: An ascorbate-deficient semi-dwarf mutant asfL-1 was detected in 250 Gy γ-ray treated grass pea (Lathyrus sativus L.) cv. BioR-231. The mutant contained only 42 % of leaf and 20 % of root ascorbate content of mother control (MC). I investigated the possible causes of ascorbate deficiency and its effect on growth and antioxidant defense in control and 150 mM NaCl-treated seedling after 60 d growth period. Ascorbate deficiency was due to significant reduction in activities of monodehydroascorbate reductase and dehydroascorbate reductase as well as increase in ascorbate oxidase, leading to considerable decrease in redox state. Despite low ascorbate pool and decrease in ascorbate peroxidase activity, shoot and root biomass production in asfL-1 mutant were similar to MC plants, even at NaCl treatment. High accumulation of glutathione (GSH) coupled with high activities of GSH reductase, catalase, GSH peroxidase and peroxidase in both tissues of the mutant permitted efficient recycling of GSH and scavenging of H2O2 through well integrated catalase/peroxidase system, despite high superoxide dismutase activity under NaCl treatment. The collapse of this system led to inhibition of growth in NaCl-treated mother plants. Together, the results suggested that asfL-1 plants undertook a major reshuffle in its antioxidant defense machinery, which effectively counterbalanced the negative impact of ascorbate deficiency and remained unperturbed by NaCl treatment to maintain normal growth and biomass production.

Photosynthetic and leaf anatomical characteristics of Castanea sativa: a comparison between in vitro and nursery plants

Abstract: The anatomic and functional leaf characteristics related to photosynthetic performance of Castanea sativa growing in vitro and in nursery were compared. The irradiance saturated photosynthesis in in vitro grown plantlets was significantly lower compared to nursery plants (65 vs. 722 μmol m−2 s−1). The maximum photosynthetic rate (PNmax) was 4.0 and 10.0 μmol(CO2) m−2 s−1 in in vitro microshoots and nursery plant leaves, respectively. Carboxylation efficiency (CE) and electron transport rate (ETR) were three-folds higher in nursery plants than in microshoots. The nonphotochemical quenching (NPQ) was saturated at 80 μmol m−2 s−1 in microshoots suggesting limited photoprotection by thermal dissipation. The microshoots had wide open, spherical stomata and higher stomatal density than nursery plants and they had almost no epicuticular wax. Consequently, the microshoots had high stomatal conductance and high transpiration rate. These anatomic and functional leaf characteristics are likely major causes of the low survival rates of plantlets after ex vitro transfer.

Effects of 5-aminolevulinic acid on the H2O2-content and antioxidative enzyme gene expression in NaCl-treated cucumber seedlings

Abstract: The potential of 5-aminolevulenic acid (ALA) to enhance the salt tolerance of cucumber (Cucumis sativus L.) seedlings was investigated. ALA was applied at various concentrations (0, 1, 10, 25, 50, and 100 mg dm−3) as foliar spray or root watering. Then the seedlings were exposed to 0 or 75 mM NaCl for 5 d. NaCl stress reduced the root and leaf dry masses, leaf area, and the leaf net CO2 assimilation rate. These reductions were counteracted by exogenous ALA, and the most efficient was 50 mg dm−3 concentration via foliar spray. ALA decreased the H2O2 contents and increased the activities of ascorbate peroxidase (APX) and glutathione reductase (GR) in NaCl-treated cucumber roots and leaves and the activity of catalase (CAT) in leaves. The ALA application also up-regulated the expressions of CAT and cAPX genes in roots and leaves and the expression of GR gene in roots of the NaCl treated cucumber plants.

Cadmium exclusion a key factor in differential Cd-resistance in Thlaspi arvense ecotypes

Abstract: Differences in Cd accumulation and Cd tolerance between Thlaspi arvense ecotype Aigues Vives (AV) from a commercial grower in South France and ecotype Jena collected in the polluted urban area of Jena (Germany) were reported here. Ecotype Jena exhibited considerable Cd-tolerance. Shoot and root masses were unaffected and root elongation was even enhanced by exposure to 50 μM Cd. In contrast, growth of ecotype AV was severely affected by this Cd treatment. Ecotype Jena was much more efficient in excluding Cd from both roots and shoots than ecotype AV. Despite the efficient restriction of Cd transport from roots to shoots in Jena, this ecotype maintained high root to shoot transport of Zn and Fe under Cd exposure. Cd supply strongly decreased the activities of antioxidant enzymes in AV, while in the Cd resistant Jena these activities either remained unaffected (SOD, APX) or were increased (CAT) by Cd supply. In conclusion, naturally selected Cd-tolerance in Thlaspi arvense is due to efficient Cd exclusion. The mechanisms underlying exclusion of Cd from the shoots seem Cd-specific yet they did not affect the homeostasis of Fe and Zn in the shoots.

Expression of the BBM gene during somatic embryogenesis of Arabidopsis thaliana

Abstract: The relationship between somatic embryogenesis (SE) and the expression of the BABY BOOM (BBM) gene was studied in cultured immature zygotic embryos (IZEs) using a transgenic line of Arabidopsis thaliana containing a BBMPro::GUS construct. Results showed spatio-temporal differences in BBM expression in explants during culture. BBM promoter activity was observed in freshly isolated IZEs except distal parts of cotyledons. At the beginning of culture, considerable increase of GUS staining intensity was observed in all parts of explants, which maintained at high level over next few days and coincide with cell divisions. Gradual decrease of GUS distribution in explants was observed at about the 5th day of culture. BBM promoter activity became largely restricted to dividing cells, then to developing somatic embryos, shoot-like structures and callus. In parts of explants not involved in morphogenesis BBM promoter activity was absent or hardly seen. Thus the in vitro expression of BBM coincides with cell proliferation and morphogenesis.

Engineering ascorbic acid biosynthetic pathway in Arabidopsis leaves by single and double gene transformation

Abstract: Six genes, which encode enzymes involved in ascorbic acid (AsA) biosynthesis, including guanosine diphosphate (GDP)-mannose pyrophosphorylase (GMP), GDP-mannose-3′,5′-epimerase (GME), GDP-galactose guanylyltransferase (GGT), L-galactose-1-phosphate phosphatase (GPP), L-galactose dehydrogenase (GDH) and L-galactono-1,4-lactone dehydrogenase (GLDH) were transformed into Arabidopsis thaliana, to evaluate the contribution of each gene to AsA accumulation. Additionally, two combinations, GGT-GPP and GGT-GLDH, were co-transformed into Arabidopsis with a reliable double-gene transformation system. AsA content of GGT transgenic lines was 2.9-fold higher as compared to the control, and co-transformation led up to 4.1-fold AsA enhancement. These results provided further evidence that GGT is the key enzyme in plant AsA biosynthesis.

Improved drought resistance in a wheat stay-green mutant tasg1 under field conditions

Abstract: We investigated the drought resistance of a wheat (Triticum aestivum L) stay-green mutant tasg1 and its wild-type (WT) in field experiments conducted for two years Drought stress was imposed by controlling irrigation and sheltering the plants from rain Compared with the WT, tasg1 exhibited a distinct delayed senescence under both normal and drought stress conditions, as indicated by slower degradation of chlorophyll and decrease in net photosynthetic rate than in WT At the same time, tasg1 mutants maintained more integrated chloroplasts and thylakoid ultrastructure than did WT plants under drought stress Lower malondialdehyde content and higher antioxidative enzyme activities in tasg1, compared to WT, may be involved in the stay-green phenotype and drought resistance of tasg1

Involvement of G6PDH in heat stress tolerance in the calli from Przewalskia tangutica and Nicotiana tabacum

Abstract: Glucose-6-phosphate dehydrogenase (G6PDH) has been implicated in supplying reduced nicotine amide cofactors for biochemical reactions and in modulating the redox state of cells. In this study, the role of G6PDH in thermotolerance of the calli from Przewalskia tangutica and tobacco (Nicotiana tabacum L.) was investigated. Results showed that Przewalskia tangutica callus was more sensitive to heat stress than tobacco callus. The activity of G6PDH and antioxidant enzymes (ascorbate peroxidase, catalase, peroxidase and superoxide dismutase) in calli from Przewalskia tangutica and tobacco increased after 40 °C treatment, although two calli exhibited a difference in the degree and timing of response to heat stress. When G6PDH was partially inhibited by glucosamine pretreatment, the antioxidant enzyme activities and thermotolerance in both calli significantly decreased. Simultaneously, the heat-induced H2O2 content and the plasma membrane NADPH oxidase activity were also reduced. Application of H2O2 increased the activity of G6PDH and antioxidant enzymes in both calli. Diphenylene iodonium, a NADPH oxidase inhibitor, counteracted heatinduced H2O2 accumulation and reduced the heat-induced activity of G6PDH and antioxidant enzymes. Moreover, exogenous H2O2 was effective in restoring the activity of G6PDH and antioxidant enzymes after glucosamine pretreatment. Western blot analysis showed that G6PDH gene expression in both calli was also stimulated by heat and H2O2, and blocked by DPI and glucosamine under heat stress. Taken together, under heat stress G6PDH promoted H2O2 accumulation via NADPH oxidase and the elevated H2O2 was involved in regulating the activity of antioxidant enzymes, which in turn facilitate to maintain the steady-state H2O2 level and protect plants from the oxidative damage.

Hydrogen peroxide and nitric oxide promote reproductive growth in Litchi chinensis

Abstract: Vegetative growth and reproductive growth strongly competes with each other during panicle development in litchi (Litchi chinensis Sonn.). We herein investigated the roles of hydrogen peroxide and nitric oxide in the competition between growth of rudimentary leaves and panicle development. The results show that the chilling-induced flowering increased H2O2 and NO contents in the mixed buds. Treatments with sodium nitroprusside (SNP), the NO donor, and methyl viologen dichloride hydrate (MV), the superoxide generator, increased NO and H2O2 contents in the mixed buds. MV and SNP treatments promoted abscission of rudimentary leaves and encouraged panicle development before or at the stage of panicle emergence. The nitric oxide synthase inhibitor Nω-nitro-L-arginine methyl ester (L-NAME) and the H2O2 trapper dimethylthiourea (DMTU) inhibited a chilling-induced flowering. SNP promoted the expression of litchi LEAFY homolog (LcLFY). These promotive effects were suppressed by the NO scavenger, 2-phenyl-4,4,5,5-tetramethylimidazoline-1-oxyl3-oxide (PTIO) and the H2O2 trapper, DMTU. The results suggest that H2O2 and NO promote reproductive growth by inhibiting the growth of rudimentary leaves as well as by promoting the expression of the flower related gene, LcLFY.

Cu/Zn superoxide dismutase activity and respective gene expression during cold acclimation and freezing stress in barley cultivars

Abstract: The transcript levels and activities of the superoxide dismutase isoenzyme (Cu/ZnSOD) were assessed in winter (Tarm-92) and spring (Zafer-160) barley cultivars during cold acclimation, freezing stress and after rewarming. Leaf Cu/ZnSOD activity and Cu/ZnSOD expression level were not significantly changed during cold acclimation. The Cu/ZnSOD expression increased evidently at mild freezing stress (−3 °C; F1), while Cu/ZnSOD1 activity did not show any response and Cu/ZnSOD2 activity decreased continuously during F1 and F2 (−7 °C) in Tarm-92. On the other hand, root Cu/ZnSOD2 activity was in accordance with Cu/ZnSOD expression in Zafer-160 after F2 treatment. Rewarming periods did not cause any significant changes in the Cu/ZnSOD activity and expression of Cu/ZnSOD in both cultivars when compared to freezing stresses. These results showed that freezing stress can regulate differently Cu/ZnSOD transcription and enzyme activity.

Changes in antioxidative enzymes activities during Tacitus bellus direct shoot organogenesis

Abstract: Changes in antioxidative enzymes activities during Tacitus bellus direct shoot organogenesis from leaf explants were examined. During the early stages of shoot organogenesis there was a decrease in superoxide dismutase (SOD) and an increase in catalase (CAT) activity, and later during organogenesis there was an increase in peroxidase (POD) and polyphenol oxidase (PPO) activity. Two highly regulated turning points may be distinguished regarding activities and isoforms of antioxidative enzymes: the initiation of shoot organogenesis and the shoot bud formation. Our data suggest the role of specific CAT, POD, SOD and PPO isoforms in separate processes during T. bellus direct shoot organogenesis.

Expression of Na + /H + antiporter gene in response to water and salinity stress in grapevine rootstocks

Abstract: Grapevine rootstocks belonging to different species of Vitis differ in their response to stress. Vines of rootstocks 110R and 1613C were subjected to different salt and water stresses individually and in combination. Expression of VvNHX1, a Na+/H+ antiporter gene, was analyzed at 7 and 21 d of stress treatment. In 110R, the expression of VvNHX1 gene increased in response to both salinity and water stress already after 7 d. Under salinity and combined stress enhanced expression of this gene was observed also after 21 d whereas expression decreased under water stress. In 1613C, expression of this gene did not increase under salinity stress. There was delayed response to water and combined stress and expression increased several fold after 21 d of stress. The stressed vines of 110R maintained lower sodium content and higher K+/Na+ ratio as compared to rootstock 1613C.

Cytosolic NADP-isocitrate dehydrogenase in Arabidopsis leaves and roots

Abstract: NADP-dependent isocitrate dehydrogenase (NADP-ICDH) catalyses the production of NADPH, which is an essential component in the cellular homeostasis. In Arabidopsis, the kinetic parameters (Km and Vmax) of cytosolic NADP-ICDH were different in leaves and roots. In vitro applied H2O2 did not affect the NADP-ICDH activity in either organ, however, the reduced glutathione inhibited the activity in leaves but not in roots. On the other hand, S-nitrosoglutathione (a NO donor) and peroxynitrite depressed NADP-ICDH activity in leaves and roots.

Sexual incompatibility in Rosaceae fruit tree species: molecular interactions and evolutionary dynamics

Abstract: Fruit crops have a growing economic importance worldwide and molecular genetics might be useful in solving many problems that arise during commercial production. One of the fields that have attracted intense attention is the molecular basis of self-incompatibility that may result in low fruit set. In tree fruits of the Rosaceae family, the incompatibility reactions take place between the pistil S-ribonuclease (S-RNase) and the pollen-expressed S-haplotype specific F-box (SFB) proteins. In most cases, the loss of self-incompatibility was associated with mutations in the S-RNase or SFB genes. A total of 27 non-functional S-haplotypes have been identified and characterized, most (24) of which emerged as a consequence of natural mutations. In the Prunoideae, most haplotypes are pollen-part mutants (50 %), while 8 are stylar-part mutants (36 %), one haplotype shows both pollen- and stylar-part mutations, and molecular changes for two haplotypes still have not been clarified. In contrast, non-functional natural haplotypes in the Maloideae are all stylar-part mutants. The analysis of such mutants may shed light on underlying molecular mechanisms as was the case with the establishment of the general inhibitor model that describes interactions between pollen and pistil S-proteins. However, several other molecules were supposed to contribute to the molecular interactions, at least in Solanaceae, a family with a similar self-incompatibility system. This review also endeavours to delineate the evolutionary implications of the S-locus mutations and collect limited data on non-S-locus molecular interactions and signaling events after self- and cross-pollination of fruit tree species.

Physiological and ultrastructural effects of lead on tobacco

Abstract: The effects of lead toxicity on leaf gas exchange, chlorophyll content, chlorophyll fluorescence, chloroplast ultrastructure, and opening of stomata were examined in tobacco (Nicotiana tabacum L.) plants. Plants were grown in a growth chamber for 7 d in Hoagland nutrient solution supplemented with 0.0 (control), 5, 10, 25, 50, 100, 300 and 500 μM Pb(NO3)2. Plants treated with 5, 10, and 25 μM Pb(NO3)2 exhibited no abnormalities. Root and shoot growth, net photosynthetic rate and stomatal conductance were significantly reduced in plants treated with 100, 300 and 500 μM Pb(NO3)2. In plants treated with 500 μM Pb(NO3)2, the majority of stomata were closed. The effect of Pb(NO3)2 on chlorophyll content and chlorophyll fluorescence parameters was negligible. However, in plants exposed to 100, 300, and 500 μM Pb(NO3)2, the mesophyll cells showed altered chloroplasts with disrupted thylakoid membranes.

Characterization of a drought inducible trithorax-like H3K4 methyltransferase from barley

Abstract: Histone H3 lysine 4 methylations catalyzed by histone lysine methyltransferases (HKMTs), like the Arabidopsis thaliana ATX1 and ATX2, are important epigenetic modifications related to chromatin decondensation and gene activation. In order to study this epigenetic mechanism in monocot cereal plants, we performed homology searches of ATX1 and ATX2 against the Brachypodium distachyon L. Beauv and rice (Oryza sativa L. spp. japonica) genomes, discovering single homologues for each cereal crop representing both Arabidopsis sequences. Using this information, we employed the rolling circle amplification — rapid amplification of cDNA ends (RCA-RACE) method to isolate, clone and characterize HvTX1 from RNA extracted from barley (Hordeum vulgare L.) tissues and studied its expression during seed development and under drought stress. The cloned cDNA sequence contained a 3 093 bp ORF homologous to ATX1 and ATX2. Characterization of the translated HvTX1 transcript sequence revealed the multi-domain nature of the putative protein, including all conserved regions characteristic for ATX1 and ATX2. By comparative genomic analysis and homology searches in EST databases we located, with high probability, the gene coding for HvTX1 on the barley chromosome 5H. Constant elevation of HvTX1 expression was observed during seed development. Expression of HvTX1 after drought stress was analyzed by quantitative real-time polymerase chain reaction (qPCR) in two different barley cultivars with varying drought stress tolerance, revealing HvTX1 drought-induction in a tolerance-specific manner.

Ectopic expression of peanut acyl carrier protein in tobacco alters fatty acid composition in the leaf and resistance to cold stress

Abstract: Acyl carrier protein (ACP), as an essential protein cofactor, plays an important role in de novo synthesis of fatty acids in plastids. In this study, the expression profile of peanut (Arachis hypogaea) AhACP1-1 and AhACP1-2 was analyzed in different tissues. The expression level of AhACP1-1 was highest in the seed, whereas expression was barely detected in the shoot, and AhACP1-2 was expressed in every tissue analyzed with the highest expression level detected in the leaf and seed. Overexpression (OE) and antisense-inhibition (AT) of AhACP1 in transgenic tobacco modified the transcript level of endogenous NtACPs, and the content of total lipids and composition of fatty acid in leaves were altered compared with the wild-type control. Transgenic OE-AhACP1 or AT-AhACP1 tobacco exhibited a significant increase or decrease in polyunsaturated C18:2 and C18:3 fatty acid content, and were more tolerant or sensitive to cold stress, respectively. It is suggested that AhACP1 bound with C18:1 might be the specific substrate of oleoyl-ACP thioesterase or glycerol-3-phosphate acyltransferase, and participates in membrane lipid synthesis.

Isolation and expression of a cold-responsive gene PtCBF in Poncirus trifoliata and isolation of citrus CBF promoters

Abstract: C-repeat/dehydration-responsive element binding factor (CBF) plays important roles in cold response network in plants. Here, one member of CBF coding gene family in trifoliate orange (Poncirus trifoliata), designated as PtCBF, was isolated. Semi-quantitative reverse transcription-polymerase chain reactions showed up-regulation of PtCBF not only under low temperature but also induced by abscisic acid. Additionally, the CBF genomic fragments in four citrus species including trifoliate orange, sweet orange (Citrus sinensis), pummel (Citrus grandis) and rough lemon (Citrus jambhiri) were isolated with complete open reading frames. According to the results of alignment analysis between full length cDNA and genomic DNA sequences in trifoliate orange, there were no introns in PtCBF. Moreover, the results of multiple sequence alignment analysis and phylogenetic analysis on putative protein sequences suggested that the AP2 DNA binding domains and CBF signature sequences were highly conserved in four citrus CBF proteins. Finally, the CBF promoters in above citrus species were isolated, which provides some information concerning promoter function.

Engineering resistance against Tobacco streak virus (TSV) in sunflower and tobacco using RNA interference

Abstract: The coat protein (CP) gene of Tobacco streak virus (TSV) from sunflower (Helianthus annuus L.) was amplified, cloned and sequenced. A 421 bp fragment of the TSV coat protein gene was amplified and a gene construct encoding the hairpin RNA (hpRNA) of the TSV-CP sequence was made in the plasmid pHANNIBAL. The construct contains sense and antisense CP sequences flanking a 742 bp spacer sequence (Pdk intron) under the control of the constitutive CaMV35S promoter. A 3.6 kb Not I fragment containing the hpRNA cassette (TSV-CP) was isolated from pHANNIBAL and sub-cloned into the binary vector pART27. This chimeric gene construct was then mobilized into Agrobacterium tumefaciens strain LBA4404 via triparental mating using pRK2013 as a helper. Sunflower (cv. Co 4) and tobacco (cv. Petit Havana) plants were transformed with A. tumefaciens strain LBA4404 harbouring the hpRNA cassette and in vitro selection was performed with kanamycin. The integration of the transgene into the genome of the transgenic lines was confirmed by PCR analysis. Infectivity assays with TSV by mechanical sap inoculation demonstrated that both the sunflower and tobacco transgenic lines exhibited resistance to TSV infection and accumulated lower levels of TSV compared with non-transformed controls.

The development of stomata and other epidermal cells on the rice leaves

Abstract: In the leaves of rice (Oryza sativa), stomatal initials arose from two asymmetric cell divisions and a symmetric division. Guard mother cells (GMCs) and long cells in stomatal files (LCSs) were formed through the first asymmetric division of the precursor cell of GMCs. Subsidiary cells (SCs) were produced by the second asymmetric division of subsidiary mother cells or LCSs. Following SC formation, GMCs divided once symmetrically to generate guard cells and then differentiated terminally to form mature stomata. The developmental patterns of long cells, prickle hairs and short cells (phellem cells and silica cells) were also examined. Interestingly, we found that the different developmental stages of stomata and epidermal cells occurred in the similar location of immature leaves of the same phyllotaxis. In addition, two spacing patterns ("one stoma, one long cell" and "one short cell row") probably exist in rice leaves.

Impact of short-term cadmium treatment on catalase and ascorbate peroxidase activities in barley root tips

Abstract: The aim of this study was to analyse the relationship between root growth inhibition and alterations in catalase (CAT) and ascorbate peroxidase (APX) contents and activities in barley root tips during the recovery after short-term Cd stress. Significant root growth inhibition was observed after a relatively short-term (30 min) exposure of barley roots to low 15 μM Cd concentration. In seedlings treated with 30 μM Cd root growth was renewed 8–9 h after Cd treatment. By contrast, seedlings exposed to 60 μM Cd failed to restore root growth. The CAT activity increased after 15 μM Cd treatment, whereas, higher Cd concentrations inhibited CAT activity. However, APX activity was not affected by Cd treatment. The content of APX1 transcript increased while content of APX2 decreased 3 h after short-term treatment in a concentration dependent manner. While the expression of CAT1 was upregulated after 15 and 30 μM Cd treatment, it was strongly downregulated by 60 μM Cd. By contrast, CAT2 was upregulated in a concentration dependent manner. These results suggest that increased CAT activity is crucial for restoration of root growth after moderate Cd stress while at severe Cd stress its inhibition may lead to the irreversible damages.

Modified alternative oxidase expression results in different reactive oxygen species contents in Arabidopsis cell culture but not in whole plants

Abstract: Alternative oxidase (AOX) transfers electrons from ubiquinone to oxygen in the respiratory chain of plant mitochondria. It is widely accepted that AOX functions as a mechanism decreasing the formation of reactive oxygen species (ROS) produced during respiratory electron transport. However, there are no experimental data to provide unambiguous proof of this hypothesis. We have studied growth characteristics, ROS content, and stress sensitivity in Arabidopsis transgenic lines with reduced or increased levels of AOX. We demonstrated that AOX-deficient plants grown in soil had an extended reproductive phase. Changes in AOX activity did not affect ROS content or stress sensitivity in the whole plants. However in the suspension culture, cells overexpressing AOX had significantly lower ROS content, whereas the AOX-deficient cells had higher ROS contents compared to the wild-type (WT) cells. Prooxidant treatment led to the increase in ROS content and to the reduction of viability more in the cells overexpressing AOX than in WT and AOX-deficient cells. Thus, we demonstrated that differences in the metabolism of whole plants and cultured cells might affect AOX functioning.

Exogenous H2O2 increased catalase and peroxidase activities and proline content in Nitraria tangutorum callus

Abstract: Antioxidative responses and proline accumulation induced by exogenous H2O2 were investigated in the callus from halophyte Nitraria tangutorum Bobr. H2O2-treated callus exhibited higher H2O2 content than untreated callus. The activities of catalase (CAT) and peroxidase (POD) significantly increased in the callus treated with H2O2, while ascorbate peroxidase (APX) activity decreased. In addition, significantly enhanced proline content was observed in the callus treated by H2O2, which could be alleviated by H2O2 scavenger dimethylthiourea and calcium (Ca) chelator ethylene glycol bis-(β-aminoethyl ether)-N,N,N′,N′-tetra-acetic acid (EGTA). Moreover, γ-glutamyl kinase (GK) activity increased in H2O2-treated callus, but proline dehydrogenase (PDH) activity decreased significantly, and the reduction was partly abolished by EGTA or Ca channel blocker verapamil. Assays using a scanning electron microscope showed significantly enhanced Ca content in H2O2-treated callus.

Overexpression of AtHsp90.3 in Arabidopsis thaliana impairs plant tolerance to heavy metal stress

Abstract: The functions of cytosolic heat shock protein AtHsp90.3 in response to heavy metal stress were characterized by using expression of AtHsp90.3 gene in yeast and Arabidopsis thaliana. AtHsp90.3 supported the Saccharomyces cerevisiae Hsp90 knockout strain R0005 growth and maintaining cells membrane integrity under cadmium and arsenic stresses, which was compatible with the components of ScHsc82 machinery. However, constitutive overexpression of AtHsp90.3 in Arabidopsis impaired plant tolerance to Cd stress with lower germination rate and shorter root length, decreased contents of phytochelatins (PCs) and glutathione (GSH), inhibited activities of superoxide dismutase (SOD), catalase (CAT) and peroxidase (POD), and increased content of malondialdehyde (MDA). These results suggested that proper homeostasis of Hsp90 was critical for cellular response and/or tolerance to heavy metal stress in plants.

Monitoring of cultivar identity in micropropagated olive plants using RAPD and ISR markers

Abstract: Randomly amplified polymorphic DNA (RAPD) and inter simple sequence repeat (ISSR) markers were applied to assess the genetic stability of micropropagated olive (Olea europaea L. cv. Maurino) plants regenerated by axillary buds. Initial olive explants, isolated from one donor tree, were multiplied on Murashige and Skoog medium for 12 repeated subcultures. A total of 40 RAPD and 10 ISSR markers resulted in 301 distinct and reproducible band classes showing homogeneous RAPD and ISSR patterns. The amplification products revealed genetic stability among the micropropagated plants and between them and the donor plant. The results demonstrate the genetic stability of nine year old mature micropropagated olive plants cultured in field, and corroborated the fact that axillary multiplication is the safest mode for multiplication of true to type plants.

Changes in protein pattern during different developmental stages of somatic embryos in chickpea

Abstract: Mature embryonic axes were used for chickpea (Cicer arietinum L.) regeneration via somatic embryogenesis. Qualitative and quantitative estimation of protein profile during somatic embryogenesis by SDS-PAGE and densitometric analysis showed differential expression of various storage proteins at different stages of somatic embryo development, which was compared with the profile of developing seeds. Total protein content in somatic embryos of chickpea increased from globular stage [2.9 μg mg−1(f.m.)] to cotyledonary stage [4.8 μg mg−1(f.m.)] and then started decreasing during onset of maturation and germination [up to 1.5 μg mg−1(f.m.)]. Differential expression of seed storage proteins, late embryogenesis abundant (LEA) proteins and proteins related with stress response were documented at different stages of somatic embryogenesis. Germinating somatic embryos showed degradation products of several seed storage proteins and the appearance of new polypeptides (76.8, 67.6, 49.9 and 34.2 kDa), which were absent during differentiation of somatic embryos. A low molecular mass (17.7 kDa) polypeptide was uniformly present during all stages of somatic embryogenesis and it may belong to a group of stress-related proteins. This study describes the expression of true seed storage proteins like legumin, vicilin, convicilin and their subunits at different stages of somatic embryogenesis, which may serve as excellent markers for embryogenic pathway of regeneration in chickpea.