Immunocytochemical analysis using antibody raised against human H2AX histones phosphorylated at serine 139 (γ-H2AX) demonstrates that root meristem cells of Vicia faba exposed to UV-radiation or incubated with hydroxyurea (HU) reveal discrete foci at the border of the nucleolus and perinucleolar chromatin or scattered over the whole area of cell nucleus. Western blots detected only one protein band at the position expected for the phosphorylated form of H2AX. The dose-effect relationship was demonstrated following treatment with 2.5 and 10 mM HU. Proteins extracted from root meristems incubated for 2 h either with HU and caffeine or with HU and sodium metavanadate showed unchanged amounts of bound γ-H2AX antibodies, as compared to root meristems treated with 2.5 mM HU. Higher quantities of phosphorylated H2AX histones were detected in proteins extracted from roots treated with HU and 2-aminopurine. All treatments were effective in producing evident aberrations of premature mitosis: broken and lagging chromatids, acentric fragments, chromosomal bridges and micronuclei. Our results show that phosphorylation of H2AX at the carboxy-terminal Ser-Gln-Glu sequence is among the earliest responses to double-strand breaks and, presumably, one of the key ATM/ATR-dependent signals indispensable for the repair of spontaneous and induced DNA damage in plant cells.

Transgenic papayas carrying the papaya ringspot virus coat protein gene were inoculated with Papaya ringspot virus. Infection was monitored by evaluating symptoms and by reverse transcription polymerase chain reaction (RT-PCR). Among eight tested transgenic lines, clone G2 was found highly resistant to virus infection during 3 years of testing. Further analysis of this clone revealed complex multicopy transgene insertion with aberrant copies. The suspected post transcriptional gene silencing was confirmed by siRNA detection. While the R0 generation of G2 transgenic papaya was found to be fully resistant to the infection, Papaya ringspot virus was able to break this resistance in subsequent generations by suppressing post-transcriptional gene silencing (PTGS).

The experiment was conducted to identify the response of three cultivars of okra [Abelmoschus esculentus (L.) Moench] to exogenous hormones [gibberellic acid-(GA₃) and prohexadione-Ca] applied as foliar spray. Stem and leaf dry masses and stem length were significantly enhanced by the application of exogenous GA₃, but prohexadione-Ca inhibited growth. Control and prohexadione-Ca treated okra plants took more time to bloom than did GA₃ treated plants. In the fruits of all the cultivars a decrease in fructose content was observed, while protein content remained almost unchanged after the application of the two growth regulators. The small changes in chlorophyll a fluorescence characteristics observed under prohexadione-Ca suggested a weakening of the photochemical processes near the photosystem 2 reaction centre. The lowering of ratio between maximum time to reach maximum fluorescence, F_m (T_max) and Area (sum of F_m-F_t for t = 0 to t = T_max) caused by GA₃ was probably due to the increase of Area rather than to changes in T_max.

The influence of different sugars (sucrose, maltose, glucose and fructose, 0.05-0.5 M) on embryogenesis and plant regeneration from cultured anthers of niger [Guizotia abyssinica (L. f.) Cass.] have been studied. Among the different sugars tested, 0.2 M sucrose was the best for embryo induction and plant regeneration. Maximum of 57 embryos per 60 anthers were induced on embryo induction medium [Gamborg's B5 medium supplemented with 10 µM 2,4-dichlorophenoxyacetic acid (2,4-D), 2 µM kinetin (KIN)] containing 0.2 M sucrose. Embryo differentiation was achieved on B5 medium supplemented with 0.5 µM benzyladenine (BA) and 0.09 M sucrose. Embryo maturation was on B5 medium containing 10 µM abscisic acid (ABA) and 0.09 M sucrose. Embryo germination was achieved on B5 medium with 0.09 M sucrose. Embryos that were developed on B5 medium supplemented with 0.2 M sucrose showed highest frequency (68 %) of plant regeneration.

The activity of antioxidant enzymes in different apricot (Prunus armeniaca L.) cultivars, resistant or susceptible to Plum pox virus (PPV), was analyzed during the years 2002 and 2003. Resistant cultivars showed higher activities of catalase (CAT), ascorbate peroxidase (APX) and dehydroascorbate reductase (DHAR) than susceptible cultivars. Only CuZn-SOD isozymes were detected in the apricot cultivars. However, no correlation was observed between this isozyme pattern and the resistance to PPV. On the other hand, PPV-resistant apricot cultivars could have a greater capability for elimination of H₂O₂ and recycling of ascorbate-glutathione cycle, and they have at least two of these enzymatic activities (CAT, APX and DHAR) over the average. In contrast, this response was not observed in the susceptible cultivars. All these data suggest that the activities of CAT, APX and DHAR could be used as biochemical markers of sharka resistance in apricot.

Sunflower plants were transformed via co-cultivation of previously bombarded hypocotyl explants with Agrobacterium tumefaciens harboring the plasmid pNGL that contains the human lysozyme gene. The transformed shoots were selected using kanamycin and regenerated plants were analyzed using histochemical β-glucuronidase assay. Southern, Western and Northern blot analyses indicated the transfer, expression and stable integration of the foreign DNA into the sunflower genome. Resistance against the phytopathogenic fungus Sclerotinia sclerotiorum, which causes white mold disease, was confirmed using a phytopathogenic test and microscopic observation of the infection process.

A 1 μM solution of ammoniates [ZnCu(NH₃)_n]²⁺(CO₃)^2- was inserted into a cut shoot of flax with the transpiration stream of water. Analysis of the ¹⁴C content after ¹⁴CO₂ assimilation by the shoot showed that ammoniates increased radioactive label contents in the tissues (especially in the young leaves and stem). In the leaves the higher sucrose to hexoses ratio, an increased radioactivity of glycerate and malate and decreased incorporation of ¹⁴C into oligosaccharides and pigments were observed. These effects were more pronounced in the young leaves. Spraying of plants with 20 mM solution resulted in an increase of plant height and leaf number.

The influence of sugars and growth regulators on shoot and root growth of Dactylorhiza species was studied under in vitro conditions. The seedling development was stimulated with the application of glucose and sucrose at concentration of 10 g dm^-3 each. The improvement of shoot growth rate and shoot length was enhanced by cytokinins N ⁶-(2-isopentenyl)adenine or N ⁶-benzyladenine and their combination with auxin indolebutyric acid (IBA). The root growth rate and root length of seedlings increased in the presence of IBA and α-naphthaleneacetic acid. Individual Dactylorhiza species showed statistically significant differences in shoot and root development depending on sugar and growth regulator combinations.

The effect of 24-epibrassinolide (BR₂₇) on cold resistance of rape seedlings was studied by ion leakage and photosynthetic pigment degradation measurements. Aqueous solutions of BR₂₇ were injected into cotyledons or primary leaves of rape plants and these plants were incubated at 2 °C or 20 °C. Cold treatment (2 °C) without BR₂₇ injection elevated the membrane permeability in both primary leaves and cotyledons significantly. Surprisingly, injection of leaves with water or 0.467 % aqueous ethanol solution led to a massive increase in membrane permeability after cold stress at 2 °C. The synergistic effect of leaf infiltration and cold on permeability was abolished by 0.05 and 1.00 µM of BR₂₇ in primary leaves and by 1.00 µM of BR₂₇ in cotyledons. On the other hand, BR₂₇ solutions strongly elevated the membrane permeability at 20 °C, while water and ethanol solutions brought about only negligible increases. Water or ethanol infiltrations strongly reduced the leaf contents of chlorophyll (Chl) a, Chl b and carotenoids at 2 °C but less markedly at 20 °C. However, in seedlings exposed to 2 °C pigments content was significantly higher in BR₂₇-treated leaves as compared to water/ethanol control. There were no differences between pigment contents of leaves injected with BR₂₇ solutions or only water/ethanol at 20 °C. The above data strongly support the stress protecting effect of BR₂₇.

The paper is focused on two groups of proteins inevitably important for cellulose biosynthesis in vascular plants. These are cellulose synthases and chitinase-like proteins. Cellulose synthases have been the subject of much research, and current conceptions and recent findings are reviewed in this paper. Severe effects of mutations and expression analysis have recently shown that chitinase-like proteins are crucial components of cellulose biosynthesis. However, understanding of their precise function is missed. Further research is to be prompted by an effective idea on it. I propose that chitinase-like proteins could play a role in the assembly of nascent glucan chains into microfibrills. Therefore, cellulose synthases and chitinase-like proteins are possibly sequential elements of the cellulose biosynthesis.

Young plants of a rhizomatous grass Calamagrostis epigejos (L.) Roth were grown from seed in nutrient solutions containing nitrogen in concentrations 0.1, 1.0, and 10 mM. After six weeks of cultivation the plants were defoliated and changes in growth parameters and in content of storage compounds were measured in the course of regrowth under highly reduced nitrogen availability. Plants grown at higher nitrogen supply before defoliation had higher amount of all types of nitrogen storage compounds (nitrates, free amino acids, soluble proteins), which was beneficial for their regrowth rate, in spite of lower content of storage saccharides. Amino acids and soluble proteins from roots and stubble bases were the most important sources of storage compounds for regrowth of the shoot. Faster growth of plants with higher N content was mediated by greater leaf area expansion and greater number of leaves. In plants with lower contents of N compounds number of green leaves decreased after defoliation significantly and senescing leaves presumably served as N source for other growing organs. Results suggest that internal N reserves can support regrowth of plants after defoliation even under fluctuating external N availability. Faster regrowth of C. epigejos with more reserves was mediated mainly by changes in plant morphogenesis.

DNA content was estimated by flow cytometry in seedlings and in vitro clones of six species: Oenothera paradoxa, Inula verbascifolia ssp. aschersoniana, Rubus chamaemorus, Solidago virgaurea, S. graminifolia and Pueraria lobata. With the exception of P. lobata, there was no difference in genome sizes between seedlings and in vitro cultured plants from any species, indicative that they maintain their genetic stability during in vitro culture. This confirms the usefulness of tissue culture for production of certified plant material to obtain herbal medicines.

The effects of 100 and 200 µM methyl jasmonate (MJA) on cell proliferation and paclitaxel and baccatin III production were investigated in free and alginate immobilized cells of Taxus baccata growing in a selected product formation culture medium. The greatest accumulation of paclitaxel (13.20 mg dm^-3) and baccatin III (4.62 mg dm^-3) occurred when 100 µM MJA was added to the culture medium of cells entrapped using a 1.5 and 2.5 % alginate solution. The effects of different treatments on the viability of cultured cells and their capacity to excrete both taxanes into the surrounding medium were considered.

The endosperms of Carthamus tinctorius cv. HUS-305, excised at globular to heart-shaped stages of zygotic embryo development, were cultured on Murashige and Skoog's medium (MS) supplemented with different concentrations of 6-benzylaminopurine (BAP), kinetin, thidiazuron (TDZ), 2,4-dichlorophenoxyacetic acid (2,4-D) or α-naphthalene-acetic acid (NAA). The highest incidence of callusing was on 2,4-D supplemented media. However, embryos differentiated only from the calli developed on media supplemented with BAP, kinetin or TDZ with the last eliciting maximum embryogenic response. The addition of a reduced nitrogen source, casein hydrolysate to MS medium supplemented with BAP and/or NAA, did not stimulate the response. However, adenine sulphate (100 mg dm^-3) promoted the induction of somatic embryos. Upon transfer to MS basal medium or the same supplemented with 0.61 µM gibberellic acid (GA₃), plumular poles of few embryos elongated resulting in the development of shoots.

Procedures were developed for flow cytometric analysis and sorting of mitotic chromosomes (flow cytogenetics) of common vetch (Vicia sativa L., 2n=12). Suspensions of intact chromosomes were prepared from root tips after cell cycle synchronization, formaldehyde fixation, and mechanical homogenization. On average, 3 × 10⁵ morphologically intact chromosomes could be isolated from 25 root tips. Flow cytometric analysis of DAPI-stained chromosomes resulted in histograms of relative fluorescence intensity (flow karyotypes) containing four peaks, representing particular chromosomes and/or pairs of chromosomes with similar relative DNA content. Peaks I and II were assigned to chromosomes 6 and 5, respectively. These chromosomes could be sorted with a purity exceeding 90 %. The two remaining peaks on the flow karyotype were composite, each of them representing a pair of chromosomes. Chromosomes 1 and 3 were assigned to composite peak III while chromosomes 2 and 4 were assigned to composite peak IV. The chromosomes could be sorted with a purity of 99 % from both composite peaks. Bivariate flow karyotyping after simultaneous staining of chromosomes with DAPI and mithramycin was not found helpful in discriminating additional chromosomes. This study extends the number of legume species for which flow cytogenetics is available and provides a new tool for targeted and effective analysis and mapping of common vetch genome.

Quantitative trait loci (QTLs) for Cu-tolerance were determined in wheat grown in control and Cu-treated soil in greenhouse. In addition, loci having an influence on the shoot Cu-, Fe-, Mn-and Zn-contents under non-stressed and Cu-stressed environments were mapped. One major QTL for Cu-tolerance was found on chromosome 5DL, while slighter effects were determined on the chromosomes 1AL, 2DS, 4AL, 5BL and 7DS. QTLs affecting the shoot Mn-and Zn-contents were found on the chromosomes 3BL and 3AL, respectively. The centromeric region on the chromosome 3B plays a role in the regulation of the shoot Fe-contents in the stressed plants. Under Cu-stress QTL affecting shoot Cu-content was found on chromosome 1BL, while on the chromosome 5AL a QTL influencing the Cu-accumulation ability of wheat from Cu-polluted soil was determined.

Changes in dry matter accumulation and allocation, abscisic acid content and carbon isotope composition of three wheat cultivars from dry, middle and wet climate regions were recorded at full maturity after exposure to different watering regimes (100, 50 and 25 % field capacity). Compared with the wet climate cultivar, the dry climate cultivar showed lower stem height, total leaf area, total dry biomass and total grain dry mass, and higher root/shoot ratio, abscisic acid content and carbon isotope composition under all watering regimes. Both water-limited treatments significantly reduced leaf growth and increased dry matter allocation into the roots leading to a significant raise of root/shoot ratio in all cultivars tested. In addition, drought affected morphological and physiological properties more in the dry climate cultivar than in the wet climate cultivar.

Histidine-containing phosphotransmitters (AHPs) transfer a phosphoryl group from membrane receptors to effectors in the nucleus. Five AHP genes have been identified in Arabidopsis. Real-time reverse transcription polymerase chain reaction (RT-PCR) was employed to quantify contents of individual transcripts in Arabidopsis leaves, roots, stems, flowers and siliques. High organ specificity of gene expression was found in AHP1, AHP2, AHP4 while expression of AHP3 and AHP5 appears more ubiquitous. We detected two AHP5 specific PCR products (AHP5 and AHP5L). Out of the five AHP5 introns, the second one was retained unspliced in the longer product (AHP5L) while AHP5 corresponded to a completely spliced AHP5 mRNA. The ratio between alternately and completely spliced AHP5 mRNAs was highest in flowers and lowest in siliques and stems. No evidence for alternative splicing in the remaining AHP transcripts was found. Two open reading frames were identified in AHP5L. The conserved part of the phosphotransfer domain remains unaffected in the longer one as the translation initiation at an intron ATG would result in a polypeptide coded by exons 3 to 6 and a short extension encoded by the intron part of the alternative open reading frame. However, the isoelectric point would shift by about 3 units towards neutral.

Transgenic rice plants in which the content of dienoic fatty acids was increased as a result of co-suppression of fatty acid desaturase were more tolerant to high temperatures than untransformed wild-type plants, as judged by growth rate and chlorophyll content. When untransformed wild-type and transgenic rice seedlings were incubated at 35 °C, seedlings of the transgenic rice lines showed approximately 1.6 and 2.1 times the growth of untransformed wild-type seedlings, as assayed by shoot and root mass, respectively. The chlorophyll content of the transgenic leaves after 9 d at 35 °C was also higher than that of wild-type rice. The maximum photochemical efficiency of photosystem 2 was also higher in transgenic plants than in wild-type plants upon high temperature stress.

A modified low copy number plant binary vector pUN has been constructed and successfully used to clone unstable DNA sequences. The vector pUN comprises of low copy number, broad host range RK2 replicon from pBin19 and of multiple cloning site (MCS) and T-DNA region, both from a pBINPLUS-derived pLV06 vector. The absence of the ColE1 replicon in the backbone of the binary vector significantly contributed to stability of hardly clonable DNA sequences and enabled their transfer into the tobacco plants through Agrobacterium-mediated transformation.

The in vitro response of sweet cherry rootstock Gisela 5 (Prunus cerasus × Prunus canescens) to increasing concentrations of NaCl (0, 50, 100 and 150 mM) in the Murashige and Skoog culture medium was studied. Induced salinity reduced growth and chlorophyll content in shoots but had no effect on water content. The increase in malondialdehyde content indicated that salinity induced oxidative stress which was accompanied with the visible symptoms of salt injury in the shoots. Antioxidant enzymes, such as superoxide dismutase, ascorbate peroxidase, peroxidase, catalase, and glutathione reductase were also significantly elevated. Although no change was observed in the Cl concentration, Na concentration of shoots significantly rose and NaCl treatments impaired K, Ca and Mg nutrition and induced imbalance in K:Na and Na:Ca ratios.

Several parameters affecting transient β-glucuronidase (GUS) expression in protocorms of Dendrobium cv. Jaquelyn Thomas were examined using bombardment technique with pActin-1D plasmid. The GUS activity in Dendrobium protocorm was not significantly affected by size of the target, type of particles, and helium gas pressure. However, the numbers of surviving tissues after bombardment were different. Transgenic orchids were established by bombardment of actively dividing protocorms with gold particles coated with pMNK1005 plasmid containing hygromycin phosphotransferase (hgh) and green fluorescent protein (gfp) genes driven by a ubiquitin promoter. A high efficiency of orchid transformation was established using three selection steps. The bombarded protocorms were screened on medium supplemented with 5 mg dm^-3 and 25 mg dm^-3 hygromycin and surviving protocorms were stringently selected on medium containing 30 mg dm^-3 hygromycin. The transformation efficiency was 19.87 % and GFP expressing protocorms were not chimeras. The integration of the transgene into genomic DNA of transgenic plantlets was confirmed by PCR and Southern blot hybridization. All but one of the transgenic lines contained multiple copies of the transgene.

At irradiances close to those representing a sunny day, red and green leaves of poinsettia (Euphorbia pulcherrima) showed only minor differences in their photosynthetic capacities despite the strong differences in their pigment composition. However, contrarily to green leaves, red leaves did not show inhibition of photosynthesis at high irradiances, because anthocyanins protected chloroplasts from photoinhibition.

Randomly amplified polymorphic DNA (RAPD) markers were used to assess genetic stability of 80 micropropagated Hagenia abyssinica plants, 40 of axillary origin and 40 of adventitious origin. The shoots were isolated from the same mother tree and micropropagated for over two years. Among the 83 RAPD primers screened, 16 gave reproducible band patterns. These 16 primers produced 115 bands for each plant. One plant from axillary origin showed two unique bands with primer OPC-11. All other plants showed identical band patterns. Generally, there was no significant difference in the shoot multiplication rate between shoots of axillary and adventitious origin. Indole-3-acetic acid (IAA) resulted in better ex vitro rooting compared to indole-3-butyric acid (IBA) and α-naphthaleneacetic acid (NAA). Non-micropropagated plants that were grown in the greenhouse for about one year were better in ex vitro rooting compared to those of juvenile material and mature tree derived micropropagated plants of the same treatment. Adventitious rooting related oxygenase gene (ARRO-1) isolated from apple (Malus domestica) was not expressed in H. abyssinica using a complementary DNA representational difference analysis fragment (cDNA RDA14) as a probe.

The effects of drought on growth, protein content, lipid peroxidation, superoxide dismutase (SOD), peroxidase (POX), catalase (CAT) and polyphenol oxidase (PPO) were studied in leaves and roots of Sesamum indicum L. cvs. Darab 14 and Yekta. Four weeks after sowing, plants were grown under soil moisture corresponding to 100, 75, 50 and 25 % field capacity for next four weeks. Fresh and dry masses, and total protein content in leaves and roots decreased obviously under drought. However, several new proteins appeared and content of some proteins was affected. Measurement of malondialdehyde content in leaves and roots showed that lipid peroxidation was lower in Yekta than in Darab 14. Severe stress increased SOD, POX, CAT and PPO activities in leaves and roots, especially in Yekta. According to the present study Yekta is more resistant to drought than Darab 14.

Withanolides-steroidal lactones, isolated from various Solanaceous plants have received considerable attention due to their potential biological activities. Five selected withanolides (withanone, withaferin A, withanolide A, withanolide B, withanolide E) were identified by HPLC-UV (DAD) - positive ion electrospray ionization mass spectroscopy in Withania somnifera (L.) Dunal cv. WSR plants and tissues cultured in vitro at different developmental phases. Cultures were established from five explants on Murashige and Skoog's medium supplemented with different plant growth regulators. Results suggest that production of withanolides is closely associated with morphological differentiation.