The root system of potato (Solanum tuberosum L. cv. Favorita) plants was treated with different O₂ and CO₂ concentrations for 35 d in aeroponic culture. Under 21 or 5 % O₂ in the root zones, the thickness of leaves and palisade parenchyma significantly increased at 3 600 μmol(CO₂) mol^-1 in the root zone, compared with CO₂ concentration 380 μmol mol^-1 or low CO₂ concentration (100 μmol mol^-1). In addition, smaller cells of palisade tissue, more intercellular air spaces and partially two layers of palisade cells were observed in the leaves with root-zone CO₂ enrichment. Furthermore, there was a significant increase in the size of chloroplasts and starch grains, and the number of starch grains per chloroplast due to elevated CO₂ only under 21 % O₂. In addition, a significant decline in the thickness of grana and the number of lamellas, but no significant differences in the number of grana per chloroplast were observed under elevated CO₂ concentration. The accumulation of starch grains in the chloroplast under elevated CO₂ concentration could change the arrangement of grana thylakoids and consequently inhibited the absorption of sun radiation and photosynthesis of potato plants.

The use of expressed sequence tag-simple sequence repeat (EST-SSR) markers might reflect the better relationship among species or cultivars than markers previously used. The first set of 30 EST-SSR was developed in hop (Humulus lupulus L.). They represent 25 gene loci with total of 1268 EST sequences. They were used for characterization of 11 hop samples and cross-amplification in Humulus japonicus Sieb. et Zucc. The number of alleles per locus ranged from two to nine. The observed and expected heterozygosities ranged from 0.182 to 0.956 and from 0.233 to 0.775, respectively. We used EST-SSR markers for cluster analysis of hop genotypes. Dendrogram well matched with genealogical and geographical data for hop genotypes.

Short-time direct and indirect effects of 25 μM Cd on the growth of dicotyledon (Phaseolus coccineus) and monocotyledon (Allium cepa) plants were investigated in the presence of inhibitors of ethylene synthesis, NADPH oxidase, and the octadecanoid pathway. Only 5 min-long action of Cd was enough for inhibition of growth in bean roots, but its recovery time was extended to several days. After 7 h treatment, Cd was significantly accumulated in bean roots, but maximum H₂O₂ accumulation was seen after 1 h. Cd-induced H₂O₂ accumulation decreased especially after addition of ethylene inhibitor silver thiosulphate (STS). Low Cd accumulation and high growth inhibition were observed also in bean leaves and in A. cepa roots. The inhibitors of the octadecanoid pathway greatly weakened the inhibitory effect of Cd in P. coccineus roots, while no significant effect was observed in A. cepa. NADPH oxidase and ethylene blockade reversed (in the case of bean plants and indirectly treated A. cepa plants) or significantly diminished Cd action. Cd-induced growth inhibition of P. coccineus leaves was also alleviated by most inhibitors of the jasmonate pathway and by STS. These results indicate that Cd may have indirect and direct effects on growth processes.

This review summarizes main difficulties involved in barley (Hordeum vulgare L.) transformation. The most commonly used procedures for genetic transformation in barley are Agrobacterium tumefaciens and particle bombardment mediated methods. While different barley cultivars are used for genetic engineering with varying sensitivity, recent improvements in regeneration and transformation techniques are described and summarized. Furthermore, some of the transformation complicating factors, in particular somaclonal variation and transgene insertion sites, are discussed in more detail.

In this study we identified and characterized a major latex lectin - designated as EtLLH - with antimicrobial activity from the succulent African milk tree Euphorbia trigona. The lectin is highly concentrated in the latex of E. trigona and appears to be composed of at least two subunits with a molecular mass of 32 kDa. EtLLH shares significant similarities to known plant lectins - ricin from Ricinus communis and agglutinin from Viscum album coloratum - which specifically bind D-galactose and N-acetyl-D-glucosamine, the major building blocks of many fungal cell walls. Antimicrobial activity assays revealed an impact of EtLLH on three phytopathogenic filamentous ascomycetes. The germination of the conidiospores and the hyphal growth of Aspergillus niger and Fusarium graminearum were severely inhibited by EtLLH already at concentrations below 0.1 mg cm^-3, while the effect on germination of the melanized conidiospores of Botrytis cinerea was less significant.

Differences among three clones of Gentiana punctata L. hairy root shoot regenerants were investigated in relation to their growth patterns, production of secondary metabolites and 2D protein profiles. Prominent differences in growth parameters were stable thus qualifying regenerant clones as true somaclones. Marked differences in protein spots were registered among the regenerant clones but not in comparison with the non-transformed control. Southern blot hybridization of regenerants showed the absence of rolA, B and C genes, initially present in the main hairy root lines. Orf13 and rolD were present and orf8 was missing in all three regenerant clones whereas orf3 was missing only in clone 2. Although lacking the three major rol genes, plants of regenerant clones retained characteristics of the hairy root phenotype.

This study was carried out to investigate the in vitro competition in tissue culture of three flax (Linum usitatissimum L.) cultivars using different distances among hypocotyl explants cultured. Hypocotyl fresh and dry masses, shoot regeneration percentage, shoot number per hypocotyl, regenerated shoot length and total chlorophyll content were examined during shoot regeneration, while plantlet height, number of roots and length of roots were recorded during rooting. With decreasing distance among explants we observed increased shoot regeneration and rooting till a certain point from where stress initiated and significant decreases in all parameters observed. Explants cultured at distance 1.0 cm were found to be at their optimum.

The elucidation of molecular mechanisms underlying the leaf development can be facilitated by the detailed anatomical study of leaf development mutants. We present an analysis of leaf anatomy and morphogenesis during early developmental stages in has mutant of Arabidopsis thaliana. The recessive has mutation affects a number of aspects in plant development, including the shape and size of both cotyledons and leaves. The earliest developmental observations suggest almost synchronous growth of the first two leaf primordia of has mutant. No significant disruption of the cell division pattern in the internal tissue is observed at the earliest stages of development, with the major anatomical difference compared to wild type primordia being the untimely maturation of mesophyll tissue cells in has mutant. At the stage of leaf blade formation, structure disruption becomes clearly evident, by irregular arrangement of the cell layers and the lack of polarity in juvenile has leaves. One distinguishing feature of the mutant leaf anatomy is the absence of mesophyll tissue differentiation. Altered has mutant leaf morphology could be at least partially accounted for by the ectopic STM activity that was found at the base of leaf primordia during early stages of leaf development in has plants.

The effect of exogenously applied nitric oxide on the heat tolerance of Chrysanthemum morifolium was investigated by applying the NO donor sodium nitroprusside (SNP). We found that the SNP partially alleviated the heat stress by slowing down the reduction of photosynthetic pigment content and net photosynthetic rate. SNP treatment also lowered the increase in the non-photochemical quenching of fluorescence and malondialdehyde content and maintained higher activities of superoxide dismutase, peroxidase, catalase and ascorbate peroxidase.

The removal of Mg²⁺ is an important step in the chlorophyll degradation pathway and extracts from senescent and presenescent Arabidopsis thaliana leaves were analyzed for Mg-dechelatase activity, using chlorophyllin, an artificial derivative of the natural substrate, chlorophyllide. The optimum temperature and pH for this reaction were determined to be at approximately 50 °C and 7.2, respectively. Mg-dechelatase activity was enhanced by addition of EDTA and inhibited by MgCl₂, HgCl₂ and reduced glutathione, indicating phenomenons such as retroinhibition by reaction products and dependence on the redox state of the mixture. Size exclusion chromatography was performed on Arabidopsis leaf extracts, and Mg-dechelatase activity was found in the fraction corresponding to molecular mass of about 42 kDa, which indicates that the Mg-dechelating compound in Arabidopsis is considerably larger than in other systems. During dark-induced senescence, the activity increased over time until reaching a maximum at day 4, and then decreased. The addition of plant growth regulators indicated that the accumulation of Mg-dechelatase was activated by ethylene and delayed by 6-benzylaminopurine.

Dark-grown seedlings of Picea abies (L) Karst. are able to accumulate the highest amounts of chlorophyll (Chl) and its precursor protochlorophyllide (Pchlide) in all Pinaceae, but calli derived from 14-d-old green cotyledons of P. abies are completely white during the cultivation in the dark. Pchlide reduction is catalysed in the dark by light-independent protochlorophyllide oxidoreductase (DPOR). This enzyme complex consists of three protein subunits ChlL, ChlN and ChlB, encoded by three plastid genes chlL, chlN and chlB. Using semiquantitative RT-PCR, we observed very low expression of chlLNB genes in dark-grown calli. It seems, that chlLNB expression and thus Chl accumulation could be modulated by light in P. abies calli cultures. This hypothesis is supported by the fact, that we observed low contents of glutamyl-tRNA reductase and Flu-like protein, which probably affected Chl biosynthetic pathway at the step of 5-aminolevulinic acid formation. ChlB subunit was not detected in dark-grown P. abies calli cultures. Our results indicated limited ability to synthesize Chl in callus during cultivation in the dark.

Methylglyoxal (MG) is a highly reactive α-oxoaldehyde, demonstrating anticancer effect on plant neoplastic tumours. In in vivo studies it was observed that MG destroyed crown gall tumours in Nicotiana tabacum produced by Agrobacterium tumefaciens, without any adverse effect on the host. The efficacy of MG in comparison to other anticancer drugs viz. cisplatin and ellagic acid in the treatment of crown gall was investigated. A slight degeneration of galls was noted in plants treated with cisplatin and ellagic acid but the plants died subsequently. With MG however, crown galls were completely cured and the plants completed their usual life cycle by flowering and producing seeds. MG inhibited the respiration of crown gall calluses suggesting that energy depletion resulted in tumour destruction.

Jatropha curcas L. (physic nut) is native to Central America and now naturalized widely in many tropical and subtropical areas. Microsatellite markers were isolated and characterized. Eleven out of 55 markers showed polymorphisms, and the allelic variation was investigated using 26 accessions of J. curcas collected from several provinces in Thailand. Each marker showed 2 to 5 alleles and the average polymorphic information content (PIC) was 0.49. Thirty four markers (62 %) were also successfully amplified in J. integerrima, J. gossypifolia and J. podagrica.

Mature pollen grain represents a highly desiccated structure with an extremely tough cell wall. Thanks to it, it resists common proteomic protocols. Instead, a robust homogenization has to be performed since proteins are needed to burst out of the cell to be included in the extracted proteome fraction. Here, a novel way of pollen homogenization employing Roche MagNA Lyser Instrument is presented, sparing time and laborious work. However, plant proteomics does not rely solely on perfect homogenization; also the choice of the extraction protocol is of key importance. The composition of the extraction buffer has a decisive influence on which proteome fraction will be extracted. Therefore the second part of our study is dedicated to the comparison of different extraction protocols with respect to subsequent proteomic analyses.

Three cultivars of tomato Pusa Ruby, Arka Vikas and Pusa Early Dwarf were subjected to osmotic stresses induced by mannitol, polyethylene glycol and NaCl in vitro. Polypeptide patterns were analyzed during each stress treatment to differentiate between tolerant and sensitive cultivars. The stresses induced more stress proteins in cv. Pusa Ruby compared to other two cultivars indicating it to be relatively osmotic stress tolerant.

In this study, we have analyzed the expression of the low oxygen inducible sucrose synthase isozyme SH1 (SUS-SH1) in the phloem of maize (Zea mays L.) infected with maize bushy stunt phytoplasma. Immunolocalization and Western blot analysis revealed several fold induction of SUS-SH1 in companion cells of phytoplasma inhabited phloem of leaf sheaths and stems. The results imply higher rates of sucrose metabolism and intensified hypoxia in the phloem.

Cucumis sativus L. seedlings were pretreated 1 h with 10 μM paraquat (PQ) and then were subjected to normal (25/18 °C) or elevated (42/38 °C) temperature to investigate whether PQ can protect plants against heat stress. Heat stress inhibited fresh and dry masses of the second leaf, root dry mass and shoot fresh mass. In leaves, the stress disintegrated membranes of 84.97 % chloroplasts and elevated contents of malondialdehyde, superoxide radical and hydrogen peroxide. In contrast, PQ pretreatment altered antioxidant activities in leaves, even after PQ was rinsed off before seedlings were exposed to different temperatures. Under heat stress, PQ pretreatment improved plant growth, decreased percentage of abnormal chloroplasts (53.03 %) and reduced contents of malonaldehyde, superoxide radical and hydrogen peroxide due to increased activities of antioxidant enzymes such as superoxide dismutase, catalase, guaiacol peroxidase, glutathione peroxidase, ascorbate peroxidase, monodehydroascorbate reductase, dehydroascorbate reductase and glutathione reductase.

Low expression of osmotically responsive genes 2 (LOS2) encodes an enolase (2-phospho-D-glycerate hydrolase, EC 4.2.1.11) that converts 2-phospho-D-glycerate (PGA) to phosphoenolpyruvate (PEP) in the glycolytic pathway in Arabidopsis. Meanwhile, it is a transcriptional activator of cold-responsive gene, negatively controlling the expression of STZ/ZAT10, a zinc finger transcriptional repressor of cold-responsive gene from Arabidopsis. A novel LOS2 gene, designated PtrLOS2 (GenBank accession number GQ144341), was isolated from trifoliate orange [Poncirus trifoliata (L.) Raf.]. The PtrLOS2 cDNA is 1 662 bp in length with a 1 338 bp open reading frame (ORF), encoding a deduced 445 amino acid residue protein with a predicted molecular mass of 47.79 kDa and an isoelectric point of 5.54. The deduced protein of the PtrLOS2 gene shares high identity (over 86 %) with other plant species enolase, which suggests that the PtrLOS2 probably encodes an enolase. Sequence alignment showed that PtrLOS2 protein has a conserved DNA-binding and a repression domain. Moreover, a conserved start site of alternative translation for the c-myc promoter binding protein (MBP-1) was also found in PtrLOS2 protein. PtrLOS2 was constitutively expressed in leaves, stems and roots. PtrLOS2 expression in roots and stems was much higher than that in leaves under normal conditions, however, the expression of PtrLOS2 was up-regulated in leaves, but down-regulated in roots after cold treatments. The PtrLOS2 expression in stems was firstly up-regulated and then down-regulated after cold treatments. Meanwhile, after ABA treatment, the expression of PtrLOS2 was up-regulated in leaves but in stems and roots firstly down-regulated followed with up-regulation.

Arabidopsis thaliana plants (wild type accessions Col and N1438) were grown in nutrient solution for 34 d with or without 50 mM NaCl. Salt stress inhibited plant growth rate more in Col than in N1438 and a decrease in K⁺, Ca²⁺ and nitrogen contents was observed in both accessions. NaCl diminished accumulation of malate, fumarate and citrate only in Col accession. To measure the effect of NaCl on transcript level of superoxide dismutase (SOD) isoforms and copper chaperone for SOD genes, a semi-quantitative polymerase chain reaction (RT-PCR) method was developed using cDNA normalized against the EF1a gene in parallel with quantitative real time RT-PCR (Q-PCR) technique. Both methods gave the same results. The abundance of transcripts of the three genes coding for Cu/Zn-SOD responded similarly to NaCl in both accessions: CSD1 gene was overexpressed, and CSD2 and CSD3 genes were repressed. However, the genes coding for Fe-SOD (FSD1), Mn-SOD (MSD1) and Cu-chaperone for SOD (CCS) responded to NaCl differently in Col and N1438: the former gene was overexpressed in Col and repressed in N1438, and the opposite behaviour was observed for the latter two genes.

The effects of two-day dark treatment, applied to whole plants or to individual organs, on the photosynthetic apparatus in cotyledons and first rosette leaves of young Arabidopsis thaliana plants were studied. Darkness affected the individually darkened pair of cotyledons as well as the cotyledons of whole darkened plants (DP) in a similar manner as revealed by the significant decrease in the actual yield of photosystem 2 electron transport and the down-regulation of the psaB and rbcL transcript levels. However, cotyledons and rosette leaves responded differently to darkness with respect to the non-photochemical quenching (NPQ) and the non-regulated energy dissipation (Φ_NO), indicating different capacity for photoprotection depending on the type of the applied dark treatment. Besides, the expression of the genes for the two plastid proteases FtsH5 and Deg1 involved in D₁ protein degradation was inhibited in both leaf organs, suggesting that these proteases function mainly under irradiance. Upon re-irradiation, dark-treated cotyledons recovered from the applied stress and during further senescence the changes in the photosynthetic parameters and the mRNA levels of psaB, rbcL and SAG12 were similar as in the control plants. However, in the course of recovery typical chloroplast senescence symptoms were observed only in individually darkened leaves while re-irradiated DP leaves maintained high photosynthetic capacity.

To determine the effects of leaf colour on gas exchange and chlorophyll fluorescence, two genotypes of Begonia semperflorens with green leaves or red leaves were compared. The red leaves showed a high accumulation of anthocyanins and high absorbance at 282 and 537 nm while the green leaves exhibited a higher net photosynthetic rate and lower thermal dissipation of light energy. It seems likely that anthocyanins in the vacuoles restricted the absorption of green light to the chloroplasts, leading to a decrease in the efficiency of excitation capture by open PS 2 centres, photochemical quenching and CO₂ assimilation.

The growth and ion content of salt sensitive Lycopersicon esculentum Mill. cv. M82 and salt tolerant L. pennellii Correll accession LA716 were examined under both control and stress conditions (150 mM NaCl). L. esculentum grew more vigorously than L. pennellii under optimal conditions, however, L. pennellii was able to maintain its growth better than cultivated tomato when the plants were exposed to salinity. Sodium content of both L. esculentum and L. pennellii increased as a result of NaCl stress. In addition, both species showed reduced potassium and calcium content due to salinity. The physiological traits were also measured in a population of 52 L. pennellii introgression lines grown under both normal and stress conditions. A total of 311 quantitative trait loci (QTL) were identified for the studied traits: plant height, stem diameter, leaf number, leaf and root fresh and dry mass, and sodium, potassium and calcium contents. Some of the loci (124) were identified under both control and stress conditions while 86 QTL were identified only under non-stress conditions and 101 loci were identified only under NaCl stress.

To broaden our knowledge of flower development, two floral homeotic genes, LbAG and LbSEP3, were isolated from the flower of Lycium barbarum L. The open reading frame length of LbAG and LbSEP3 were 1090 and 992 bp encoding 249 and 242 amino acids, respectively. Sequence alignment and phylogenetic analysis indicated that LbAG belonged to C-type MADS-box gene and that LbSEP3 was E-type MADS-box gene. Compared with other floral homeotic proteins, LbAG held the conserved AG motif I, II and LbSEP3 conserved SEP3 motif I, II. Expression profile showed that LbAG transcripts were abundant in inner two whorls and fruit but not in root, leaf, sepal, and petal, and that LbSEP3 constitutively expressed in root, leaf, fruit, and all the four floral whorls.

Sour pummelo (Citrus grandis) seedlings were irrigated with nutrient solution containing four boron concentrations (i.e., 2.5, 10, 25 and 50 μM H₃BO₃) and two aluminum concentrations [i.e., 0 (-Al) and 1.2 mM AlCl₃ . 6 H₂O (+Al)]. It was found that B did not affect, but Al increased, the Al content in the roots. The Al and citrate contents in the -Al leaves either did not change or slightly increased with increasing B concentration. On the other hand, the Al and citrate contents in the +Al leaves rapidly decreased as B concentration increased from 2.5 to 50 μM, then decreased at the highest B concentration. The Al and citrate contents were higher in the +Al than in the -Al leaves, except for at 25 μM B when they were similar. The leaf malate content did not change in response to B or Al, except for an increase in the +Al leaves and a decrease in the -Al leaves at 2.5 μM B. Similarly, the root malate and citrate contents did not change in response to B with or without Al, except for a decrease in the malate and citrate contents in the +Al roots at 50 μM B and an increase in the citrate content in the -Al roots at 50 μM B. The activities of acid-metabolizing enzymes were less affected by B-Al interactions in the roots than in the leaves.

Changes in pigments contents, leaf area, leaf dry mass per unit area (LMA), photosynthetic rate and chlorophyll a fluorescence were investigated in developing leaves of Fagopyrum dibotrys Hara. mutant. Anthocyanins transiently accumulate below the upper epidermis during leaf ontogeny of this mutant. Red leaves possessed lower Chl content, LMA, photosynthetic rate and apparent carboxylation efficiency than green leaves. However, content of anthocyanins declined and above mentioned parameters increased during further leaf development. In both red and green leaves, chronic photoinhibition did not take place according to variable to maximum chlorophyll fluorescence ratio (F_v/F_m). Red leaves had higher non-photochemical quenching (NPQ) and higher PS 2 efficiency.

Metallothioneins are involved in detoxification of heavy metals. A cDNA encoding type 3 metallothionein (PcMT3) was isolated from the salt stressed leaf cDNA library of Porteresia coarctata (Roxb.) Tateoka (wild rice) that grows well in the heavy metal laden estuarine soils. The PcMT3 cDNA (581 bp) encodes a protein of 64 amino acids. PcMT3 is highly homologous (82 %) to OsMT-I-3a of rice, but is unique from other type 3 plant MTs due to the presence of an additional glycine residue in the C-terminal domain. Analysis of the 5' upstream region of PcMT3 showed the presence of cis-acting elements like the CG box and STRE previously reported to be involved in gene expression under heavy metal stress. Southern analysis suggested the presence of more than one copy of PcMT3-like sequences in the P. coarctata genome. Analysis of genomic clone of PcMT3 revealed the presence of two introns. A comparison of the genomic sequence of PcMT3 with closely similar type 3 MTs from rice and mangrove species revealed conservation in the number and position of introns. Transcript profiling for PcMT3 in P. coarctata leaves in the presence of Cd, Cu and Zn showed an increase in transcript accumulation.

Universal (consensus) primers are those primers that have the ability to amplify the targeted region of DNA across a broad range of individuals in a certain group of organisms. In plants, such universal primers have been designed to target regions in the nuclear, mitochondrial or chloroplast genome. Among these three genomes, the chloroplast genome is the most suited for the design of consensus primers due to the lower rate of evolution and hence conservation of gene order and sequence of the genome among the different plant species compared to the other two genomes. Several molecular studies in plants have developed and used chloroplast-specific universal primers. In this review, I present some examples of the nuclear DNA-specific universal primers and discuss the features of the chloroplast DNA that make it the most suited for the design of such primers. I then refer to all chloroplast-specific primers developed so far and provide some examples of molecular studies and applications that made use of them.

Grape (Vitis vinifera L.) seedlings grown in vitro were treated with either 200 mM NaCl or 350 mM mannitol for 7 d. Both salinity and osmotic stress caused significant increase in electrolyte leakage. From the three commonly occurring free polyamines (PA), only conspicuous accumulation of putrescine was found in the NaCl-treated seedlings. Four PA biosynthetic genes encoding arginine decarboxylase (pVvADC), S-adenosylmethionine decarboxylase (pVvSAMDC), spermidine synthase (pVvSPDS) and spermine synthase (pVvSPMS) were successfully isolated. While induction of pVvADC was observed from the 1^st day of salt treatment, pVvSAMDC and pVvSPMS were induced only at late stage of stress. As for expression levels of genes in the mannitol-treated seedling, either temporary (pVvADC at day 1) or late (pVvSPMS at days 5 and 7) induction was observed.

Callus of the halophyte Mesembryanthemum crystallinum was used to study the effect of NaCl on the response to Botrytis cinerea infection. The fungus easily colonized the callus surface and the intercellular spaces. However, in the NaCl-adapted tissues the incidence of penetration was 67 % lower than in the inoculated control tissue. The modification of the infection pattern found in the salt-adapted callus could be related to metabolic adaptations to salinity. This was manifested by the enhanced antioxidant potential of ascorbate, the up-regulated activities of ascorbate peroxidase, as well as guaiacol and syringaldazine peroxidases together with the increased detoxification capacity of glutathione transferase in the NaCl-adapted callus. The post-inoculation changes in NaCl-adapted and non-adapted calli were roughly similar and supported the prooxidative nature of B. cinerea infection.

We investigated whether changes in abscisic acid (ABA) content in leaves of Mikania micrantha infected by the holoparasite Cuscuta campestris at five growth stages, influenced the host stomatal conductance (g_s), transpiration rate (E) and net photosynthetic rate (P_N). C. campestris infection caused a negative effect on g_s, E and P_N of the host plants. ABA content in host leaves infected by C. campestris was significantly lower at 6 d after parasitization (DAP) and significantly higher at 13 and 33 DAP, relative to uninfected controls. In the parasite, ABA content was lowest at 13 DAP and then sharply increased to the maximum at 26 DAP. Moreover, the ABA content in the parasite was always lower than in the infected host leaves. The results suggest that an increase in host ABA concentration contributes to reduced host g_s, E and P_N in the holoparasitic C. campestris - M. micrantha association.