Marcela Gubišová

Rhizofiltration potential of Arundo donax for cadmium and zinc removal from contaminated wastewater

The main aim of the presented work was to design and characterize the rhizofiltration system comprised of giant reed plants (Arundo donax L.), as species with phytoremediation potential, for the removal of cadmium as a toxic metal and zinc as an important microelement from model solutions spiked with radionuclides 109Cd and 65Zn. The possibility of effective cadmium and zinc rhizofiltration by the root system of giant reed plants under continuous flow conditions to achieve complete decontamination was confirmed. The highest 109Cd and 65Zn specific radioactivity (Bq g−1; dry mass) was found in case of plants located near the input of the solution into the system, whereby the radioactivity in the plants decreased in the direction of the solution flow within the system. On the other hand, gradual increasing of the concentration ratio [Cd]shoot: [Cd]root within this horizontal plants location was observed. Individual experiments showed that the efficiency of cadmium rhizofiltration decreased in the order of model solutions: synthetic wastewater (χ = 505 μS cm−1), deionized water (χ = 29.6 μS cm−1), 100 % Hoagland medium (χ = 1675 μS cm−1). The same results were also found in case of zinc rhizofiltration. For the description of cadmium and zinc rhizofiltration processes, the kinetic mathematical models widely applied for sorption processes under continuous flow conditions were successfully used.

Biosynthesis of Essential Polyunsaturated Fatty Acids in Wheat Triggered by Expression of Artificial Gene

The artificial gene D6D encoding the enzyme ∆6desaturase was designed and synthesized using the sequence of the same gene from the fungus Thamnidium elegans. The original start codon was replaced by the signal sequence derived from the wheat gene for high-molecular-weight glutenin subunit and the codon usage was completely changed for optimal expression in wheat. Synthesized artificial D6D gene was delivered into plants of the spring wheat line CY-45 and the gene itself, as well as transcribed D6D mRNA were confirmed in plants of T0 and T1 generations. The desired product of the wheat genetic modification by artificial D6D gene was the γ-linolenic acid. Its presence was confirmed in mature grains of transgenic wheat plants in the amount 0.04%–0.32% (v/v) of the total amount of fatty acids. Both newly synthesized γ-linolenic acid and stearidonic acid have been detected also in leaves, stems, roots, awns, paleas, rachillas, and immature grains of the T1 generation as well as in immature and mature grains of the T2 generation. Contents of γ-linolenic acid and stearidonic acid varied in range 0%–1.40% (v/v) and 0%–1.53% (v/v) from the total amount of fatty acids, respectively. This approach has opened the pathway of desaturation of fatty acids and production of essential polyunsaturated fatty acids in wheat.

Elicitation Phenolic Compounds in Cell Culture of Vitis vinifera L. by Phaeomoniella chlamydospora

Abstract The in vitro cell cultures of Vitis vinifera L. cv. St. Laurent were treated with two elicitors - synthetic methyl jasmonate and natural, prepared from grapevine plant infected with the Phaeomoniella chlamydospora, the agent causing the Esca disease of grapevine. Efficiency of phenolic compounds production after elicitation of cell culture was analysed immediately after treatment (15 min, 30 min, 60 min) and later (after 24, 48, and 72 hours). The cell growth and content of phenolic compounds (+)-catechin, (-)-epicatechin, p-coumaric acid, syringaldehyde, rutin, vanillic acid, and trans-resveratrol were analysed in cultivated cells as well as in cultivation medium. Pch-treatment increased production of total polyphenols the most significantly 15 min after the elicitation and in optimal time was 2.86 times higher than in nonelicited culture and 1.44 times higher than in MeJa induced cell culture.

A new high-molecular-weight glutenin subunit from the slovak wheat (Triticum aestivum L.) cultivar ‘Trebišovská 76’

Historical and local varieties are an important source of genetic diversity in connection with the wheat baking quality. By analysis of DNA from the coding part of the genome of the Slovak regional wheat (Triticum aestivum L.) variety ‘Trebišovská 76’, differences in the Dy subunit composition coded by locus Glu-1D were found. Comparing the newly found sequence with a generally used 1Dy12 (GenBank accession no. X03041) sequence, existence of a new high-molecular-weight glutenin subunit, which was marked as 1Dy12* (GenBank accession no. EU266533) was confirmed. The molecular weight of probable protein coded by 1Dy12* subunit reaches 72.35 kDa comparing to the 1Dy12, which reaches 68.71 kDa. This difference is not likely to distinguish by SDS-PAGE. The easiest way for cereal genotyping according to baking quality is proteomic analysis. More exact and suitable method for genotype identification appears to be marker assisted selection using PCR.

Optimization of Barley Mature Embryo Regeneration and Comparison with Immature Embryos of Local Cultivars

Optimization of Barley Mature Embryo Regeneration and Comparison with Immature Embryos of Local Cultivars Regeneration ability of plant cells or tissues in explant culture is one of the key factors affecting success of genetic transformation. In experiments, the effect of explant type (whole embryo, scutellum embryonic axis, meristematic/central zone of embryonic axis) and plant growth regulators (BAP or TDZ) on mature embryo regeneration was determined. Explant type significantly affected regeneration efficiency. While no regenerants were observed using mature scutella, whole embryos or embryonic axes produced the highest number of regenerants. Using embryonic axes with discarded apical and basal parts, regeneration efficiency dramatically decreased. No statistical differences in regeneration were observed between BAP and TDZ added to the regeneration medium in concentration 0.1 or 1 mg l-1. At last, regeneration ability of mature embryos of nine Slovak spring barley cultivars (Donaris, Ezer, Levan, Ludan, Nitran, Pribina Sladar, Orbit, Pax) and Golden Promise as a model cultivar was examined and compared with regeneration ability of immature embryos which have been usually used for genetic transformation of barley. Although the regeneration from mature embryos was very weak, the same cultivars Golden Promise, Pribina and Levan showed the best regeneration ability by using both, immature and mature embryos. On the other hand cultivars Ezer and Pax belonged to the weakest ones in both experiments.

Imaging of photoassimilates transport in plant tissues by positron emission tomography

Abstract The current findings show that positron emission tomography (PET), primarily developed for medical diagnostic imaging, can be applied in plant studies to analyze the transport and allocation of wide range of compounds labelled with positronemitting radioisotopes. This work is focused on PET analysis of the uptake and transport of 2-deoxy-2-fluoro[18F]-D-glucose (2-[18F]FDG), as a model of photoassimilates, in tissues of giant reed (Arundo donax L. var. versicolor) as a potential energy crop. The absorption of 2-[18F]FDG and its subsequent transport in plant tissues were evaluated in both acropetal and basipetal direction as well. Visualization and quantification of the uptake and transport of 2-[18F]FDG in plants immersed with the root system into a 2-[18F]FDG solution revealed a significant accumulation of 18F radioactivity in the roots. The transport rate in plants was increased in the order of plant exposure through: stem > mechanically damaged root system > intact root system. PET analysis in basipetal direction, when the plant was immersed into the 2-[18F]FDG solution with the cut area of the leaf of whole plant, showed minimal translocation of 2-[18F]FDG into the other plant parts. The PET results were verified by measuring the accumulated radioactivity of 18F by direct gamma-spectrometry.

Progress in the genetic engineering of cereals to produce essential polyunsaturated fatty acids

Essential polyunsaturated fatty acids with more than two double bonds and length of carbon chain 18-22 must be taken in the diet to prevent diseases and imbalances caused by their deficiency. Terrestrial sources of polyunsaturated fatty acids are limited to only a few plant species whose large-scale cultivation is not possible and the production of their seeds and oil is ineffective. The complete biosynthetic pathway of fatty acids is known in organisms, including plants. After the first gene encoding the enzyme catalysing the initial steps of PUFA biosynthesis (ω-3 desaturase, Δ6-desaturase) were isolated, isolation of other genes encoding relevant enzymes of the PUFA pathway from different donor organisms followed. Genetic transformations of model plants by the desaturase- and elongase-encoding genes opened the way for the genetic engineering of oilseed crop species. Some of the developed transgenic plants produced PUFAs, including eicosapentaenoic and docosahexaenoic acids. Seed oils extracted from them were similar to fish oil. Tools of the synthetic biology can be applied in modifications of the PUFA pathway and also in overcoming of limitations when the gene and its expression product are absent in the pathway. Such progress in cereals (barley, wheat, maize) has been made only recently, when the first successful modifications of the ω-3 and ω-6 PUFA pathways succeeded. This review focuses on genetic modifications of the PUFA biosynthetic pathway in cereals in relation to the status reached in model plants and oilseed crops.

Biomass Production of Gigantic Grasses Arundo donax and Miscanthus × Giganteus in the Dependence on Plant Multiplication Method

Abstract The effect of plant propagation method on growth parameters and the yield of above-ground biomass in two species of gigantic grasses were measured during three growing seasons. Plants were multiplied in explant culture and through traditional methods – by rhizome segments (Miscanthus × giganteus) or by stem cuttings (Arundo donax). In the case of M. × giganteus, in vitro-multiplied plants produced more shoots with significantly lower diameter, but the differences in the number of shoots, plant height and the yield of dry biomass were not statistically significant. Different results were observed for A. donax, where in vitro-multiplied plants showed significantly weaker results in all parameters, with the exception of the number of shoots in the first measured season. In both the species, there was observed the strong effect of the year. While in M. × giganteus the yield of dry biomass gradually decreased during the measured years, it increased in the case of giant reed.

Comparison Of Cd And Zn Accumulation In Tissues Of Different Vascular Plants: A Radiometric Study

Abstract The aim of the present work was to compare the accumulation and translocation of Cd and Zn in plants of tobacco (Nicotiana tabacum L.), celery (Apium graveolens L.), maize (Zea mays L.), giant reed (Arundo donax L.), and alpine pennycress (Noccaea caerulescens L.) under conditions of short-term hydroponic experiments using nutrient solutions spiked with radionuclides 109Cd or 65Zn, and direct gamma-spectrometry. It was found that the time-course of metals accumulation in studied plants was not different in terms of target metal, but it was significantly different on the level of plant species. The highest values of Cd accumulation showed plants of giant reed, whereby the accumulation decreased in the order: giant reed > tobacco > alpine pennycress >> maize and celery. On the basis of concentration ratios (CR) [Me]shoot / [Me]root calculation for both metals, it was found that Cd and Zn were in prevailing part accumulated in the root tissues and only partially accumulated in the shoots, where the amount of accumulated Cd and Zn increased from the oldest developed leaves to the youngest developed leaves. The CR values corresponding to these facts were calculated in the range 0.06 – 0.27 for Cd and for Zn 0.06 – 0.48. In terms of plant species, the CR values obtained for Cd decreased in the order: maize > celery > tobacco and giant reed > alpine pennycress. The similarity between studied objects – individual plant species on the basis of the obtained variables defining Cd or Zn accumulation at different conditions of the experiments as well as the relationships between obtained variables and conditions of the experiments were subjected to multivariate analysis method – cluster analysis (CA). According to the findings and this analysis, it can be expected that plants of tobacco and giant reed will dispose with similar characteristics as plants of alpine pennycress, which are classified as Zn/Cd hyperaccumulators, in terms of Cd or Zn accumulation and other positive parameters for their utilization in phytoremediation processes and techniques.

Bacterial Communities in Rhizosphere of Maize Studied by T-RFLP

Abstract The terminal restriction fragment length polymorphism munities from different collecting places was evaluated was used to determine the bacterial diversity in rhizo- by principal component analysis. Results showed that sphere of maize (Zea mays L.) collected from four sites the most different bacterial community originated from of experimental field plot in two dates of the vegetation marginal part of the experimental field plot collected in season (July and September). The 16S rRNA gene was September was caused probably by combination of the amplified from metagenomic DNA using universal eubac- marginal effect and drought before sampling date in Sep- terial primers and PCR products were digested separately tember. Other rhizosphere samples showed from moderate with three restriction enzymes. Significant differences in to small differences in the structure of the bacterial com- the number of terminal restriction fragments among rhi- munity. Nevertheless, significant differences among all zosphere samples and between sampling dates were not collected bacterial communities were not observed. detected (P < 0.05). Variation within the bacterial communities from different collecting places was evaluated by principal component analysis. Results showed that the most different bacterial community originated from marginal part of the experimental field plot collected in September was caused probably by combination of the marginal effect and drought before sampling date in September. Other rhizosphere samples showed from moderate to small differences in the structure of the bacterial community. Nevertheless, significant differences among all collected bacterial communities were not observed.