An <Emphasis Type="Italic">in vitro</Emphasis> method to evaluate grapevine cultivars for <Emphasis Type="Italic">Erysiphe necator</Emphasis> susceptibility

Powdery mildew, caused by the obligate biotrophic ascomycete Erysiphe necator, is one of the most destructive grapevine diseases worldwide. Cultivars of Vitis vinifera L, for wine and table grape production, are all susceptible to E. necator, whose attacks result in severe epidemics under the warm and dry conditions of the Mediterranean basin. The aim of the present study was to compare the susceptibility of different grapevine cultivars to E. necator by an in vitro assay for assessing the potentiality of this method in breeding programs for resistance to the pathogen. Leaves of 12 grapevine cultivars were spot-inoculated in vitro with about 10 conidia from five different isolates of E. necator, using colony growth and conidiation 3 wk post-inoculation as indicators of susceptibility to the disease. A remarkable difference was observed between highly susceptible cultivars like ‘Baresana’, ‘Malvasia’, ‘Bianca’, and ‘Italia’, and the less susceptible ‘Alphonse Lavallée’ and ‘Ohanez’, in accordance with their behavior in the field. No statistically significant differences were found in the virulence of E. necator isolates.     

Effect of <Emphasis Type="Italic">Medicago sativa</Emphasis> ferritin gene on stress tolerance in transgenic grapevine

The effect of Medicago sativa (alfalfa) ferritin gene (MsFer) on abiotic stress tolerance was tested using transgenic Vitis berlandieri × Vitis rupestris cv. ‘Richter 110’ grapevine rootstock lines. Leaf discs from transgenic plants maintained higher photosynthetic activity after NaCl, tert-butyl-hydroperoxide (t-BHP) or paraquat treatment than control ones. These results indicate that the increased production of ferritin significantly improved abiotic stress tolerance in transgenic grapevine plants.     

Up-regulation of cyclic electron flow and down-regulation of linear electron flow in antisense-<Emphasis Type="Italic">rca</Emphasis> mutant rice

To investigate how excess excitation energy is dissipated in a ribulose-1,5-bisphospate carboxylase/oxygenase activase antisense transgenic rice with net photosynthetic rate (P _N) half of that of wild type parent, we measured the response curve of P _N to intercellular CO₂ concentration (C _i), electron transport rate (ETR), quantum yield of open photosystem 2 (PS2) reaction centres under irradiation (F_v′/F_m′), efficiency of total PS2 centres (Φ_PS2), photochemical (q_P) and non-photochemical quenching (NPQ), post-irradiation transient increase in chlorophyll (Chl) fluorescence (PITICF), and P700⁺ re-reduction. Carboxylation efficiency dependence on C _i, ETR at saturation irradiance, and F_v′/F_m′, Φ_PS2, and q_P under the irradiation were significantly lower in the mutant. However, NPQ, energy-dependent quenching (q_E), PITICF, and P700⁺ re-reduction were significantly higher in the mutant. Hence the mutant down-regulates linear ETR and stimulates cyclic electron flow around PS1, which may generate the ΔpH to support NPQ and q_E for dissipation of excess excitation energy.     

<Emphasis Type="Italic">Rpv10</Emphasis>: a new locus from the Asian <Emphasis Type="Italic">Vitis</Emphasis> gene pool for pyramiding downy mildew resistance loci in grapevine

A population derived from a cross between grapevine breeding strain Gf.Ga-52-42 and cultivar ‘Solaris’ consisting of 265 F1-individuals was genetically mapped using SSR markers and screened for downy mildew resistance. Quantitative trait locus (QTL) analysis revealed two strong QTLs on linkage groups (LGs) 18 and 09. The locus on LG 18 was found to be identical with the previously described locus Rpv3 and is transmitted by Gf.Ga-52-42. ‘Solaris’ transmitted the resistance-related locus on LG 09 explaining up to 50% of the phenotypic variation in the population. This downy mildew resistance locus is named Rpv10 for resistance to Plasmopara viticola. Rpv10 was initially introgressed from Vitis amurensis, a wild species of the Asian Vitis gene pool. The one-LOD supported confidence interval of the QTL spans a section of 2.1 centi Morgan (cM) corresponding to 314 kb in the reference genome PN40024 (12x). Eight resistance gene analogues (RGAs) of the NBS–LRR type and additional resistance-linked genes are located in this region of PN40024. The F1 sub-population which contains the Rpv3 as well as the Rpv10 locus showed a significantly higher degree of resistance, indicating additive effects by pyramiding of resistance loci. Possibilities for using the resistance locus Rpv10 in a grapevine breeding programme are discussed. Furthermore, the marker data revealed ‘Severnyi’ × ‘Muscat Ottonel’ as the true parentage for the male parent of ‘Solaris’.     

Photoacclimation in <Emphasis Type="Italic">Dunaliella tertiolecta</Emphasis> reveals a unique NPQ pattern upon exposure to irradiance

Highly time-resolved photoacclimation patterns of the chlorophyte microalga Dunaliella tertiolecta during exposure to an off–on–off (block) light pattern of saturating photon flux, and to a regime of consecutive increasing light intensities are presented. Non-photochemical quenching (NPQ) mechanisms unexpectedly responded with an initial decrease during dark–light transitions. NPQ values started to rise after light exposure of approximately 4 min. State-transitions, measured as a change of PSII:PSI fluorescence emission at 77 K, did not contribute to early NPQ oscillations. Addition of the uncoupler CCCP, however, caused a rapid increase in fluorescence and showed the significance of qE for NPQ. Partitioning of the quantum efficiencies showed that constitutive NPQ was (a) higher than qE-driven NPQ and (b) responded to light treatment within seconds, suggesting an active role of constitutive NPQ in variable energy dissipation, although it is thought to contribute statically to NPQ. The PSII connectivity parameter p correlated well with F′, F _m ′ and NPQ during the early phase of the dark–light transients in sub-saturating light, suggesting a plastic energy distribution pattern within energetically connected PSII centres. In consecutive increasing photon flux experiments, correlations were weaker during the second light increment. Changes in connectivity can present an early photoresponse that are reflected in fluorescence signals and NPQ and might be responsive to the short-term acclimation state, and/or to the actinic photon flux.     

Diurnal cycle of chlorophyll fluorescence in <Emphasis Type="Italic">Phalaenopsis</Emphasis>

Chlorophyll (Chl) fluorescence of warm day/cool night temperature exposed Phalaenopsis plants was measured hourly during 48 h to study the simultaneous temperature and irradiance response of the photosynthetic physiology. The daily pattern of fluorescence kinetics showed abrupt changes of photochemical quenching (q_P), non-photochemical quenching (NPQ) and quantum yield of photosystem II electron transport (Φ_PSII) upon transition from day to night and vice versa. During the day, the course of Φ_PSII and NPQ was related to the air temperature pattern, while maximum quantum efficiency of PSII photochemistry (F_v/F_m) revealed a rather light dependent response. Information on these daily dynamics in fluorescence kinetics is important with respect to meaningful data collection and interpretation.     

Analysis of <Emphasis Type="Italic">Malus</Emphasis><Emphasis Type="Italic">S</Emphasis>-RNase gene diversity based on a comparative study of old and modern apple cultivars and European wild apple

Malus S-RNase genetic diversity was analyzed in Malus × domestica cultivars and compared to European wild apple (Malus sylvestris). Using PCR-based approaches, the S-RNase genotype of 140 M. × domestica cultivars, 196 M. sylvestris trees and 27 M. sylvestris—M. × domestica hybrids was determined. S-RNase allelic richness in M. sylvestris was much higher than in M. × domestica, indicating the negative influence of domestication on S-RNase diversity. Heterogeneity of the S-RNase allelic distribution is much higher in cultivated apple than in wild apple, which shows that breeding leads to strong departure from the expected homogeneity of genes under negative frequency-dependent selection. The majority of the M. × domestica S-alleles has been found in M. sylvestris as well, which points to strong conservation of the S-locus gene structure. Based on the sequence of all different SCAR-fragments, which comprise both the hypervariable PS1 region and the single intron, S-RNase genetic diversity was further explored. It provided some clues to the occurrence of new S-alleles among the multitude of novel S-RNase sequences that have been identified, which were mostly unique for the group of M. sylvestris individuals. The determination of the S-RNase genotypes of old cultivars and M. sylvestris will enable their introduction into new breeding strategies. As M. sylvestris has become an endangered species in Belgium, the knowledge gathered in this study will be an important tool for selecting useful genotypes for a core collection.     

Chlorophyll fluorescence in micropropagated <Emphasis Type="Italic">Rhododendron ponticum</Emphasis> subsp. <Emphasis Type="Italic">baeticum</Emphasis> plants in response to different irradiances

The aim of this study was to investigate acclimation of micropropagated plants of Rhododendron ponticum subsp. baeticum to different irradiances and recovery after exposure to high irradiance. Plants grown under high (HL) or intermediate (IL) irradiances displayed higher values of maximum electron transport rate (ETR_max) and light saturation coefficient (E_k) than plants grown under low irradiance (LL). The capacity of tolerance to photoinhibition (as assessed by the response of photochemical quenching, q_p) varied as follows: HL > IL > LL. Thermal energy dissipation (q_N) was also affected by growth irradiance, with higher saturating values being observed in HL plants. Light-response curves suggested a gradual replacement of q_p by q_N with increasing irradiance. Following exposure to irradiance higher than 1500 μmol m⁻² s⁻¹, a prolonged reduction of the maximal photochemical efficiency of PS 2 (F_v/F_m) was observed in LL plants, indicating the occurrence of chronic photoinhibition. In contrary, the decrease in F_v/F_m was quickly reverted in HL plants, pointing to a reversible photoinhibition.     

Acaricidal effects of <Emphasis Type="Italic">Corymbia citriodora</Emphasis> oil containing <Emphasis Type="Italic">para</Emphasis>-menthane-3,8-diol against nymphs of <Emphasis Type="Italic">Ixodes ricinus</Emphasis> (Acari: Ixodidae)

The toxicity of para-menthane-3,8-diol (PMD), the main arthropod-repellent compound in the oil of the lemon eucalyptus, Corymbia citriodora, was evaluated against nymphs of Ixodes ricinus using five methods (A–E) of a contact toxicity bioassay. Mortality rates were estimated by recording numbers of dead nymphs at 30 min intervals during the first 5 h after the start of exposure and at longer intervals thereafter. The mortality rate increased with increasing concentration of PMD and duration of exposure with a distinct effect after 3.5 h. From the results obtained by methods A, C and E, the LC₅₀ range was 0.035–0.037 mg PMD/cm² and the LC₉₅ range was 0.095–0.097 mg PMD/cm² at 4 h of exposure; the LT₅₀ range was 2.1–2.8 h and the LT₉₅ range was 3.9–4.2 h at 0.1 mg PMD/cm². To determine the duration of toxic activity of PMD, different concentrations (0.002, 0.01, 0.1 mg PMD/cm²) were tested and mortality was recorded at each concentration after 1 h; thereafter new ticks were tested. This test revealed that the lethal activity of PMD remained for 24 h but appeared absent after 48 h. The overall results show that PMD is toxic to nymphs of I. ricinus and may be useful for tick control.     

Genetic relatedness among some wild cherry (<Emphasis Type="Italic">Prunus</Emphasis> subgenus <Emphasis Type="Italic">Cerasus</Emphasis>) genotypes native to Iran assayed by morphological traits and random amplified polymorphic DNA analysis

Subgenus Cerasus species are useful genetic resources for cherry breeding programs. A total of 17 morphological traits together with 19 random amplified polymorphic DNA (RAPD) primers were used to study 39 accessions including 34 wild Cerasus subgenus genotypes belonging to Prunus avium L., P. cerasus L., P. mahaleb L., P. microcarpa Boiss., P. incana Pall., and P. brachypetala Boiss. species, along with an unknown wild Cerasus sample, two advanced cherry cultivars (‘Lambert’ and ‘Bulgar’), and two rootstocks (‘Colt’ and ‘Gisela 6’). Genotypes were separated into different groups according to their species and collection sites using cluster analysis performed by Ward’s clustering method based on morphological data. Nineteen RAPD primers from 60 screened produced 304 polymorphic reproducible bands (98.15% polymorphism). According to the similarity matrix, the lowest similarity was obtained between P. avium and P. microcarpa samples. A dendrogram was prepared by the unweighted pair-group method with arithmetic average (UPGMA), and the accessions were separated according to their species and geographic origin. In both morphological and molecular results, the advanced cultivars and rootstocks were separated from wild genotypes, and the unknown genotype was grouped with P. mahaleb accessions. Grouping by morphological characteristics was compared with the results of RAPD analysis, with no significant correlations between morphological and molecular data being found. This is the first report of molecular (RAPD) genetic diversity study in wild Cerasus subgenus genotypes from Iran, and the results demonstrate the high potential of RAPD analysis for discrimination of Cerasus subgenus genotypes.     

Cloning and sequence diversity analysis of <Emphasis Type="Italic">GmHs1</Emphasis><Superscript><Emphasis Type="Italic">pro-1</Emphasis></Superscript> in Chinese domesticated and wild soybeans

Soybean cyst nematode (SCN) (Heterodera glycines Ichinohe) is the most important pathogen in soybean production worldwide and causes substantial yield losses. An apparent narrow genetic base of SCN resistance was observed in current elite soybean cultivars, and searching for novel SCN resistance genes as well as novel resistance sources rather than focusing on the two important genes rhg1 and Rhg4 has become another major objective in soybean research. In the present paper we report a 1,477 bp Hs1 ^pro-1 homolog, named GmHs1 ^pro-1 . This gene was cloned from soybean variety Wenfeng 7 based on information for Hs1 ^pro-1 , a beet cyst nematode resistance gene in sugar beet. It has two domains, Hs1pro-1_N and Hs1pro-1_C, both of which are believed to confer resistance to nematodes. Of the 1,477 bp sequence in GmHs1 ^pro-1 , an open reading frame of 1,314 bp, encoding a protein with 437 amino acids, was flanked by a 5′-untranslated region of 27 bp and a 3′-untranslated region of 135 bp. Fourteen single-nucleotide polymorphisms (SNPs) were observed in 44 soybean accessions including 23 wild soybeans, 8 landraces, and 13 soybean varieties (or lines), among which 5 in wild soybeans and 3 in landrace accessions were unique. Sequence diversity analysis on the 44 soybean accessions showed π = 0.00168 and θ = 0.00218 for GmHs1 ^pro-1 ; landraces had the highest diversity, followed by wild soybeans, with varieties (or lines) having the lowest. Although we did not detect a significant effect of selection on GmHs1 ^pro-1 in the three populations, sequence diversity, unique SNPs, and phylogenetic analysis indicated a slight domestication bottleneck and an intensive selection bottleneck. High sequence diversity, more unique SNPs, and broader representation across the phylogenetic tree in wild soybeans and landraces indicated that wild collections and landrace accessions are invaluable germplasm for broadening the genetic base of elite soybean varieties resistant to SCN. C. Yuan and G. Zhou contributed to this paper equally.     

Contrasting population genetic structure and gene flow between <Emphasis Type="Italic">Oryza rufipogon</Emphasis> and <Emphasis Type="Italic">Oryza nivara</Emphasis>

The cross compatible wild relatives of crops have furnished valuable genes for crop improvement. Understanding the genetics of these wild species may enhance their further use in breeding. In this study, sequence variation of the nuclear Lhs1 gene was used to investigate the population genetic structure and gene flow of Oryza rufipogon and O. nivara, two wild species most closely related to O. sativa. The two species diverge markedly in life history and mating system, with O. rufipogon being perennial and outcrossing and O. nivara being annual and predominantly inbreeding. Based on sequence data from 105 plants representing 11 wild populations covering the entire geographic range of these wild species, we detected significantly higher nucleotide variation in O. rufipogon than in O. nivara at both the population and species levels. At the population level the diversity in O. rufipogon (Hd = 0.712; θ _sil = 0.0017) is 2–3 folds higher than that in O. nivara (Hd = 0.306; θ _sil = 0.0005). AMOVA partitioning indicated that genetic differentiation among O. nivara populations (78.2%) was much higher than that among O. rufipogon populations (52.3%). The different level of genetic diversity and contrasting population genetic structure between O. rufipogon and O. nivara might be explained by their distinct life histories and mating systems. Our simulation using IM models demonstrated significant gene flow from O. nivara to O. rufipogon, indicating a directional introgression from the annual and selfing species into the perennial and outcrossing species. The ongoing introgression has played an important role in shaping current patterns of genetic diversity of these two wild species. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Low-temperature limitation of primary photosynthetic processes in Antarctic lichens <Emphasis Type="Italic">Umbilicaria antarctica</Emphasis> and <Emphasis Type="Italic">Xanthoria elegans</Emphasis>

Temperature response curves of chlorophyll a fluorescence parameters were used to assess minimum sub-zero temperature assuring functioning of photosynthetic photochemical processes in photosystem II (PS II) of Antarctic lichens. Umbilicaria Antarctica and Xanthoria elegans were measured within the temperature range from −20 to +10°C by a fluorometric imaging system. For potential (F _V/F _M) and actual (Φ _II) quantum yields of photochemical processes the minimum temperature was found to be between −10 and −20°C. Non-photochemical quenching (NPQ) of absorbed excitation energy increased with temperature drop reaching maximum NPQ at −15°C. Image analysis revealed intrathalline heterogeneity of chlorophyll a fluorescence parameters with temperature drop. Temperature response of Φ _II exhibited an S-curve with pronounced intrathalline differences in X. elegans. The same relation was linear with only limited intrathalline difference in U. antarctica. The results showed that Antarctic lichen species were well adapted to sub-zero temperatures and capable of performing primary photosynthesis at −15°C.     

Efficient separation and purification of allophycocyanin from <Emphasis Type="Italic">Spirulina</Emphasis> (<Emphasis Type="Italic">Arthrospira</Emphasis>) <Emphasis Type="Italic">platensis</Emphasis>

Allophycocyanin (APC) is a minor component of phycobiliproteins in cyanobacteria and red algae. This paper describes a simple and inexpensive extracting method for isolating APC from Spirulina (Arthrospira) platensis with high efficiency. The crude phycobiliprotein extract was pretreated by ammonium sulfate fractionation. Then, by adding hydroxylapatite into crude phycobiliprotein extract dissolved in 20 mM phosphate buffer (pH 7.0), APC was selectively adsorbed by hydroxylapatite but C-phycocyanin (C-PC) was not. The hydroxylapatite was collected and APC was extracted from the crude phycobiliprotein extract. Then, the enriched APC was washed off from the hydroxylapatite using 100 mM phosphate buffer (pH 7.0). In this simple extracting method it was easy to remove C-PC and isolate APC in large amounts. The absorbance ratio A ₆₅₀/A ₂₈₀ of extracted APC reached 2.0. The recovery yield was 70%, representing 4.61 mg · g⁻¹ wet weight. The extracted APC could be further purified by a simple anion-exchange chromatography with a pH gradient from 5.6 to 4.0. The absorbance ratio A ₆₅₀/A ₂₈₀ of the purified APC reached 5.0, and the overall recovery yield was 43%, representing 2.83 mg · g⁻¹ wet weight. Its purity was confirmed by native polyacrylamide gel electrophoresis (PAGE) and sodium dodecyl sulfate-PAGE.     

Characterization of osmolyte betaine synthesizing sarcosine dimethylglycine <Emphasis Type="Italic">N</Emphasis>-methyltransferase from <Emphasis Type="Italic">Methanohalophilus portucalensis</Emphasis>

To overcome the extracellular salt stress, Methanohalophilus portucalensis FDF1^T synthesizes the compatible solute betaine through the methylation of glycine, sarcosine, and N,N-dimethylglycine. S-adenosylmethionine (AdoMet) is the methyl donor. The enzyme sarcosine dimethylglycine N-methyltransferase (SDMT) of M. portucalensis, that catalyzes the formation of N,N-dimethylglycine and glycine betaine, has been purified and characterized. SDMT, a monomer of 33 kDa with a pI at 5.03, has a narrow substrate specificity limited to using only sarcosine and dimethylglycine as substrates for the methyl transferase reaction. The K _m values for sarcosine and AdoMet were 2.29 and 0.21 mM, respectively, with a V _max of 0.83 μmol/mg-min (k _cat value of 0.44 s⁻¹). The K _m values for dimethylglycine and AdoMet were 3.76 and 0.59 mM, respectively, with a V _max of 4.88 μmol/mg-min (k _cat of 2.68 s⁻¹). A high concentration of the end product betaine (2.0 M) did not affect the SMT activity, but it slightly inhibited the DMT activity. Both activities were also not affected by potassium or sodium ions in concentrations of 200–1,000 mM. We compared this novel archaeal SDMT enzyme to other similar bacterial transferases as well as to the glycine sarcosine dimethylglycine methyltransferase found also in M. portucalensis.     

The tyrosine <Emphasis Type="Italic">O</Emphasis>-prenyltransferase SirD catalyzes <Emphasis Type="Italic">O</Emphasis>-, <Emphasis Type="Italic">N</Emphasis>-, and <Emphasis Type="Italic">C</Emphasis>-prenylations

Recently, the prenyltransferase SirD was found to be responsible for the O-prenylation of tyrosine in the biosynthesis of sirodesmin PL in Leptosphaeria maculans. In this study, the behavior of SirD towards phenylalanine/tyrosine and tryptophan derivatives was investigated. Product formation has been observed with 12 of 19 phenylalanine/tyrosine derivatives. It was shown that the alanine structure attached to the benzene ring and an electron donor, e.g., OH or NH₂, at its para-position are essential for the enzyme activity. Modifications were possible both at the side chain and the benzene ring. Enzyme products from seven phenylalanine/tyrosine derivatives were isolated and characterized by MS and NMR analyses including HSQC and HMBC and proven to be O- or N-prenylated derivatives at position C4 of the benzene rings. K _M values of six selected derivatives were found in the range of 0.10–0.68 mM. Catalytic efficiencies (K _cat/K _M ) were determined in the range of 430–1,110 s⁻¹·M⁻¹ with l-tyrosine as the best substrate. In addition, 7 of 14 tested tryptophan analogs were also accepted by SirD and converted to C7-prenylated derivatives, which was confirmed by comparison with products obtained from enzyme assays using a 7-dimethylallyltryptophan synthase 7-DMATS from Aspergillus fumigatus.     

Tissue culture of <Emphasis Type="Italic">Sinningia speciosa</Emphasis> and analysis of the <Emphasis Type="Italic">in vitro</Emphasis>-generated <Emphasis Type="Italic">tricussate whorled phyllotaxis</Emphasis> (<Emphasis Type="Italic">twp</Emphasis>) variant

Two protocols were developed for the efficient regeneration of Sinningia speciosa from leaf explants via two developmental pathways. The first method involved formation of callus and buds, followed by subsequent root growth, in Murashige and Skoog medium (MS) containing 2.0 mg l⁻¹ 6-benzylaminopurine (BA) and 0.2 mg l⁻¹ α-naphthalene acetic acid (NAA), with a regeneration efficiency of 99.0%. The second method involved producing callus and roots, followed by subsequent formation of buds, in MS medium supplemented with 1.0–5.0 mg l⁻¹ NAA, and resulted in a regeneration efficiency of 90.4%. Our experiments indicate that the root-first pathway resulted in a lower plant regeneration efficiency. Through five continual generations using the buds-first method, a total of 215 regenerated plants were obtained in the last generation, and eight exhibited a phenotype we named tricussate whorled phyllotaxis (twp). Six of the regenerated twp variant plants maintained their tricussate whorled phyllotaxis phenotype, showing no other abnormalities, while one reverted to a wild type before flowering and another formed two rounds of sepals. Physiological analysis revealed that the twp plants responded differently than wild type to exogenous NAA and 2,3,5-triiodobenzoic acid (TIBA), while high-performance liquid chromatography (HPLC) analysis showed that the levels of endogenous indole-3-acetic acid (IAA) and gibberellin (GA) were lower in twp than wild-type plants. These results suggest that the formation of the twp mutant may be related to phytohormones and that the twp variant could be an important material for investigating the molecular mechanism of plant phyllotaxis patterning.     

<Emphasis Type="Italic">Agrobacterium</Emphasis><Emphasis Type="Italic">tumefaciens</Emphasis>-mediated transformation of callus cells of <Emphasis Type="Italic">Crataegus</Emphasis><Emphasis Type="Italic">aronia</Emphasis>

A genetic transformation system has been developed for callus cells of Crataegus aronia using Agrobacterium tumefaciens. Callus culture was established from internodal stem segments incubated on Murashige and Skoog (MS) medium supplemented with 5 mg l⁻¹ Indole-3-butyric acid (IBA) and 0.5 mg l⁻¹ 6-benzyladenine (BA). In order to optimize the callus culture system with respect to callus growth and coloration, different types and concentrations of plant growth regulators were tested. Results indicated that the best average fresh weight of red colored callus was obtained on MS medium supplemented with 2 mg l⁻¹ 2,4-dichlorophenoxyacetic acid (2,4-D) and 1.5 mg l⁻¹ kinetin (Kin) (callus maintenance medium). Callus cells were co-cultivated with Agrobacterium harboring the binary plasmid pCAMBIA1302 carrying the mgfp5 and hygromycin phosphotransferase (hptII) genes conferring green fluorescent protein (GFP) activity and hygromycin resistance, respectively. Putative transgenic calli were obtained 4 weeks after incubation of the co-cultivated explants onto maintenance medium supplemented with 50 mg l⁻¹ hygromycin. Molecular analysis confirmed the integration of the transgenes in transformed callus. To our knowledge, this is the first time to report an Agrobacterium-mediated transformation system in Crataegus aronia.     

Expression,subcellular localization,and <Emphasis Type="Italic">cis</Emphasis>-regulatory structure of duplicated phytoene synthase genes in melon (<Emphasis Type="Italic">Cucumis melo</Emphasis> L.)

The emission of molecular iodine (I₂) from the stipe, the meristematic area and the distal blade of the brown macroalga Laminaria digitata (Hudson) Lamouroux (Phaeophyceae) was monitored under low light and dark conditions. Photosynthetic parameters were determined to investigate both the extent of stress experienced by different thallus parts and the effects of emersion on photosynthesis. Immediately after air exposure, intense I₂ emission was detectable from all thallus parts. I₂ emission declined continuously over a period of 180 min following the initial burst, but was not affected by the light regime. The total number of mole of I₂ emitted by stipes was approximately 10 times higher than those emitted from other thallus parts. Initial I₂ emission rates (measured within 30 min of exposure to air) were highest for stipes (median values: 2,999 and 5,222 pmol g⁻¹ dw min⁻¹ in low light and dark, respectively) and lower, by one order of magnitude, for meristematic regions and distal blades. After exposure to air for between 60 and 180 min, I₂ emission rates of all thallus parts were reduced by 70–80%. Air exposure resulted in a decrease of the maximum photosystem II (PSII) efficiency (F _v/F _m) by 3%, and in a 25–55% increase of the effective PSII quantum efficiency ( \Updelta F/F^￠_\textm \Updelta F/F^{\prime}_{\text{m}} ); this was caused by a higher fraction of open reaction centres (q_P), whereas the efficiency of the latter in capturing energy ( F^￠_\textv /F^￠_\textm F^{\prime}_{\text{v}} /F^{\prime}_{\text{m}} ) remained constant. The results indicate the presence of an iodine pool which is easily volatilised and depleted due to air exposure, even under apparently low stress conditions.     

Injured and uninjured leaf photosynthetic responses after mechanical injury on <Emphasis Type="Italic">Nerium oleander</Emphasis> leaves,and <Emphasis Type="Italic">Danaus plexippus</Emphasis> herbivory on <Emphasis Type="Italic">Asclepias curassavica</Emphasis> leaves

Insect herbivory has variable effects on plant physiology; so greater understanding is needed about how injury alters photosynthesis on individual injured and uninjured leaves. Gas exchange and light-adapted leaf chlorophyll fluorescence measurements were collected from uninjured and mechanical partial leaf defoliation in two experiments with Nerium oleander (Apocynaceae) leaves, and one experiment with Danaus plexippus herbivory on Asclepias curassavica (Asclepiadaceae) leaves. Gas exchange impairment (lower photosynthetic rate (P _n ), stomatal conductance (g _s)) indicates water stress in a leaf, suggests stomatal limitations causing injury P _n impairment. The same pattern of gas exchange impairment also occurred on uninjured leaves opposite from injured leaves in both N. oleander experiments. This is an interesting result because photosynthetic impairment is rarely reported on injured leaves near injured leaves. No photosynthetic changes occurred in uninjured A. curassavica leaves opposite from D. plexippus-fed leaves. Partially defoliated leaves that had P _n and g _s reductions lacked any significant changes in intercellular leaf [CO₂], C _i. These results neither support, nor are sufficient to reject, stomatal limitations to photosynthesis. Manually imposed midrib vein severance in N. oleander experiment #1 significantly increased leaf C _i, indicating mesophyll limitations to photosynthesis. Maximal light-adapted leaf photochemical efficiency () and also non-photochemical quenching (q _N) were reduced by mechanical or insect herbivory to both study species, suggesting leaf trouble handling excess light energy not used for photochemistry. Midrib injury on N. oleander leaves and D. plexippus herbivory on A. curassavica leaves also reduced effective quantum yield (ΦPSII) and photochemical quenching (q _P); so reduced plastoquinone pools could lead to additional PSII reaction center closure.