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Optimization of RNA isolation from stone fruits at different ripening stages
101-104.Views:184This study was conducted to select the most appropriate RNA isolation method that can be used successfully in case of stone fruits. The changing pattern of gene expression during the ripening process of stone fruits may elucidate the molecular background of several phenotypical or phytochemical alterations present among different genotypes. Our laboratory aims to study the expression of genes encoding for enzymes that catalyze crucial steps in the flavonoid biosynthesis pathway. RNA isolation from fruit mesocarp is a challanging task due to high levels of sugars and polyphenolics accumulating during fruit development. Therefore, at first, the optimal techniques eligible for RNA isolation from fruit tissues at different ripening stages must be selected. Our study compares three different RNA isolation protocols and describes their potential applicability according to different fruit species and ripening stages.
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Molecular analysis of strawberry cultivars using RAPD, AP-PCR and STS markers
24-28.Views:121Seventeen strawberry (Fragaria x ananassa Duch.) cultivars representing the national list of Hungary, were subjected to RAPD, AP—PCR and STS analysis. Of the 31 decamer and oligomer primers tested 26 primers produced polymorphic patterns. 45 polymorphic fragments were analysed, ranging between 200-2800 by in size. Based on the data, similarity coefficients (Jaccard index and Simple matching coefficient) were calculated, and dendrograms were constructed using the unweighted pair group method of arithmetic averages (UPGMA). The dendrograms only partly reflect the known pedigree data. Specific RAPD markers were identified for cultivars F5, Pocahontas and Rabunda.
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Shoot induction and plant regeneration from cotyledon segments of the muskmelon variety "hógolyó"
61-64.Views:138Cotyledonary segments of the casaba type muskmelon variety "Hógolyó" were used to induce organogenesis. Fifty different hormone combinations were applied to enhance the induction of shoot formation on the edge of the segments. The phases of organogenesis were followed with light- and scanning electron microscope. Shoot induction was achieved with high frequency. The shoots were transferred to hormone free media for root induction. The rooted plantlets were planted out to soil.
NAA was feasible and the method can be applied in transformation experiments.
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Fire blight (Erwinia amylovora) susceptibility of old Hungarian apple cultivars
35-38.Views:139The aim of the Hungarian apple breeding program started in the Department of Fruit Science was to find resistant apple cultivars against major diseases (scab, powdery mildew, fire blight). The outbreak of fire blight (Erwinia amylovora) in 1996 motivated us to search new resistant sources principally from old traditional apple cultivars. First of all, cultivars have been gathered since 1997 from Carpathia (Visk) and evaluated between 2001 and 2003. In this recent study evaluation of resistance of old Hungarian genotypes to fire blight collected from the English National Fruit Collection (Brogdale, Faversham) is presented.
13 old Hungarian apple cultivars in 2002, and 38 genotypes in 2005 have been evaluated. We used `Idared' and 'Jonathan M41' as susceptible controls and 'Liberty' and 'Remo' as resistant ones. Shoots of two-year-old potted plants were inoculated with a mixture of virulent E. amylovora isolates (Ea2, Ea60, Ea67) at a concentration of 5 x 108 cells/ml. Resistance of apple cultivars was evaluated weekly, four times after inoculation by disease severity of symptoms. Numbers of bacterial colonies in 1 cm length shoot were determined in the fourth week after infection.
8 cultivars in 2002 and 9 cultivars in 2005 displayed notable resistance to fire blight based on one-year data. Based on the coincident data of both years, out of the cultivars collected also from Carpathia ‘Pónyik', and `Sikulai' were found to be resistant and gene sources additional old Hungarian valuable apple genotypes could be selected: `Szabadkai szercsika' and `Tordai piros !davit'. The cultivar `Szemes alma' originated from Visk has proved to be recurrently resistant.
The number of bacterial cells in shoots of the investigated cultivars correlated with the severity of symptoms. With this method, which was introduced by us earlier, we can screen cultivars displaying weak visible symptoms, which cannot be proposed as a source of resistance because of their latent infection.
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Microsporogenesis of peach (Prunus persica L. Batsch) varieties
7-10.Views:213Bud dormancy during winter is a critical factor in peach production in Hungary. The yield is determined basically by the survival rate of flower buds during winter frosts and by their ability to develop normal floral organs. It is important to investigate the genetic basis of slow floral development during dormancy for the purpose of breeding peach varieties with better winter hardiness. The aim of the present research was to examine microsporogenesis in 14 peach varieties during three successive winters in a Hungarian germplasm collection and to study the effectiveness of this method in variety evaluation. There were significant differences in the dynamics of microsporogenesis both between the varieties and between the years. Of the varieties, ‘Mayfire', bred in California, possessed the quickest pollen development rate. The microsporogenesis of `Piroska', a Hungarian local variety, was the slowest. Rapid floral bud development was observed in `Aranycsillag', `Springcrest' and 'Venus'. A medium developmental rate was characteristic of `Babygold 6', Fairlane', `Michelini' and `Red June', while development was slow in 'Champion', 'Early Redhaven', `Redhaven', `Harko' and `Mariska'. Based on these results, the study of microsporogenesis represents a reliable method for the phenological description of peach varieties during dormancy. The application of this method makes it possible to identify varieties and landraces with slow flower bud development, suggesting better winter hardiness.
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Sensory analysis as a supporting method for marjoram breeding
73-77.Views:184Marjoram is one of the most important spices in Hungary, which is utilized both in phytotherapy as well as in alimentary industry. Organoleptic quality of the drugs of two Hungarian varieties of Majorana hortensis /'Francia' and 'Magyar'/ were investigated by two different methods of sensory analysis. The chemical composition of the material was checked by GC analysis of the distilled essential oil. Although their main compounds and proportions showed similar patterns, both the human and instrumental sensory tests based on the complex odour and aroma of the drug, proved a significant differentiation of the two varieties.
On the basis of our results, sensory evaluation may be a useful tool in the practice in determination of complex aromatic values of marjoram, as a spice. At the same time it seems to be a suitable method in promotion of breeding efforts.
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Recent developments in biochemical characterization of Vitis vinifera L. varieties in Hungary
57-61.Views:149Isoelectric focusing is an effective and well reproducible method to provide information for identification of various plant species and clones if breeding or other genetic modification(s) for a given species are reflected in changes of an isozyme pattern. The method has been used for characterization of plant proteins and enzymes and for identification of various species and varieties. Our aim was to continue our several-year-work carried out on a wide variety of grapevine varieties and to reveal whether analyses of esterase and peroxidase isozyme patterns are suitable to distinguish various grapevine varieties. Therefore we compared esterase and peroxidase isozyme patterns of various species. Plant samples were obtained from Szigetcsep, Kecskemet, Tokaj and Eger. The following samples were analyzed: Pinot gris, noir, blanc, Chardonnay, Riesling Theses, Chasselas from Szigetcs6p, Bianca and his parents Eger2 and Bouvier from Kecskemet, Furmint and Hárslevelű from Tokaj, Kékfrankos and Zweigelt from Eger. To identify various species according to their esterase isozyme patterns the after blooming phenological phase while according to their peroxidase isozyme pattern the dormant phenological phase was found as optimal sampling time.
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In vitro multiplication and hardening of grapevine plants in aeriated media
15-18.Views:205In vitro cultures have widely been used in horticulture for rapid multiplication of new varieties and clones as well as to produce pathogen-free stock material. To improve efficient hardening and transfer in vitro grown grapevine plants were multiplied by cutting them into single-node internodes with the whole leaf. Microcuttings including the shoot tips were rooted in granulated perlite moisted with tapwater under sterile conditions. After 2-3 weeks the rooted microcuttings were supplied by nutrients and hardened by gradual opening and finally by complete removal of the lids of jars or plastic boxes used for growth. Using this method microcuttings of Vitis vinifera cvs. „Chardonnay", „Cabernet franc", „Riesling" and „Sauvignon blanc" and the rootstock varieties Vitis riparia x Vitis cinerea cv. „Barrier" and Vitis berlandieri x Vitis rupestris cv. „Richter 110" formed new roots and shoots and 100% of the tested plants survived the acclimatization procedure. Similar results were obtained when perlite was replaced with rockwool-, or pit-pot blocks. This method may highly increase the efficiency of producing pathogen-free propagating material and new transgenic lines.
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Attempting Regeneration from Cultured Cotyledons and Plant Regeneration from Cotyledonary Nodes in Common Bean (Phaseolus vulgaris L.)
57-60.Views:152Dry seeds from two cultivars of common bean (Phaseolus vulgaris L.) were germinated on sterile cotton and sterile deionized distilled water. Cotyledonary node tissue of seedlings were cultured on Murashige and Skoog(MS)-based media supplemented with different combination of N6-benzyl-aminopurine (BAP) and indole-3-acetic acid (IAA), and benzyladenine (BA) and a-naphthaleneacetic acid (NAA). The results revealed that the regeneration percent and the average number of buds and shoots per explant were influenced by the type of explants and exogeneously added hormones. Multiple shoot induction on dry bean cotyledonary node that contain 4-5 mm from cotyledons and hypocotyl on a medium containing full concentration of MS inorganic salts supplemented with 0.5mg/1 BA and 0.1mg/1 NAA was feasible and the method can be applied in transformation experiments.
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Callus induction on standard type Cymbidium cultivars
108-110.Views:118Tissue cultured Cymbidium PLBs (protocormlike body) were used as starting material to induce embryogenic callus which could serve as objects of genetic transformation. We obtained callus using two methods. The first method was culturing the PLB segments for one month in liquid MS medium in the presence of 0.5 mg/1 benzyladenine and 0.05 mg/1 naphtylacetic acid followed by cultivation on the same composition solid medium with 0.5 g/l activated charcoal for an additional month. Callus formation was observed on 30% of the explants. The second way was to propagate the PLB segments on solid MS medium supplemented with 1 mg/1 thidiazuron. In these cultures we also observed callus formation on 20% of the explants.
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High-velocity microprojectile mediated DNA delivery into Phaseolus vulgaris callus cells
99-102.Views:113We report the method for the establishment of rapidly growing callus cultures of Phaseolus vulgaris and the conditions required for efficient transformation using high velocity microprojectiles and high level of transient gene expression. Using hypocotyl explant and vertical culture on B5 medium with lmg/1 kinetin and 2 mg/1 2,4-D, we can recommend to get a rapidly growing callus from bean which is a good starting material to introduce foreign DNA into bean cells. The GeneBooster particle delivery system was used for the bombardment of bean callus and the Hgm resistance gene (Hgmr) was used as a selectable marker gene. 25mg/I hygromycin (Hgm) concentration was sufficient to kill the control callus. We used the standard physical factors, the appropriate pressure of N2 gas for the bombardment of the callus tissue, the shooting distance and the size of tungsten particles used as microprojectiles. Selective and nonselective tests were made by transferring the healthy green and white calluses, subcultured for 4 months on selective and nonselective medium. Several Hgm resistant calli had been obtained. Selective pressure was maintained over a period of 10 months.
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In vitro regeneration from cotyledons of watermelon
96-98.Views:162Cotyledonary segments of five different genotypes of watermelon were used to induce organogenesis. Five different hormone combinations were applied to enhance the induction of shoot formation on the surface of the segments. The phases of organogenesis were followed with light and scanning electron microscope. Shoots were obtained after four weeks, then the shoots were transferred to hormone free medium for root induction.
This method of regeneration can be applied in transformation experiments. GUS histochemical assay was made to check the expected success of using Agrobacterium for the transformation.
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Examinations of 600-year-old seeds by means of archaeobotanical and genetical methods
79-80.Views:124About 600-year-old plant seeds were discovered in a well of a mediaeval cellar in the course of an excavation in Budapest. After the archaeobotanic purification seed of 16 species were found in large quantities. Seeds preserved in the best state were selected from each group. The existence of endosperm was analysed in these subfossils, which turned to be successful mainly in the case of grapes (Vitis vinifera) and cornels (Cornus mas). Seeds of these two species contained the most endosperm and remains of the embryo. DNA was extracted with the help of DNEasy Plant Mini Kit and analysed by RAPD-PCR method. The amplification of DNA extracted from cornel seeds resulted in detecting a 1500 by fragment, which makes the comparison of these samples with present-day cornels possible.
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Investigation of flowering dynamics of the basil (Ocimum basilicum L.) and its consequences in production
74-80.Views:157The flowering dynamics of Ocimum basilicum L., (a common population maintained at the UHFI) was studied with the aim to create an exact and practically applicable method for definition of phenological phases.
According to our observation the development of individual flowers of the basil can be characterized by 8 distinguishable phases, which must be considered for description of the actual phenological stage of a spike.
An accurate model was created for the unambiguous description of flowering process of different flowers within the spike and for the individual plant as a whole. The new flowering index formula is calculated from the number of flowers weighed by their phenological phases.
The time dependence of flowering is presented by functions fitted to values of the flowering index. The results reflected different patterns of the main inflorescence, the inflorescences formed on the side shoots in the first or in the second half of the flowering period. However, for description of the flowering process of the whole plant, a sigmoid function proved to be the appropriate model.
The accumulation process of the essential oil could be characterized by the flowering index values. They showed close correlation (r=0.964) at high probability.
The new method assures an exact definition of phenological phases in basil. Its application seems to be optimal during breeding procedures, seed production and even in production of high quality drugs, according to the requirements of the Good Agricultural Practice (Guidelines for GAP, 1998).
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Identification of plant taxons by isoelectric focusing
65-67.Views:157Differences were demonstrated in esterasei coenzyme pattern of some essential oil producing plants belonging to the Apiaceae family — fennel (Foeniculum vulgare Mill.), angelica (Angelica archangelica L.), lovage (Levisticum officinale Koch.), dill (Anethum graveolens L.), coriander (Coriandrum sativum L.), anise (Pimpinella anisum L.), caraway (Carum carvi L.) — as well as differences between two varieties of fennel seed by using isoelectric focusing. That method provides quality control in essential oil plants and is suitable to describe isoenzyme pattern characteristic for taxons.
Based on our findings, isoelectric focusing seems to be suitable for identification and differentiation of different plant samples, providing an easy tool for further processing as well as for breeding.
Our further aim is to apply that method to differentiate among samples belonging to the same species according to their value of inner content.
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Rootstocks for Cherries from the Department of Fruit Science Budapest
63-66.Views:160Cherry rootstock breeding started at the Department of Fruit Science, SSU Budapest by the late 50-s and the activity can be divided into three main groups. In the first stage the activity was focused on collection of native mahaleb cherry (Prunus mahaleb L.) varieties lead by L. Sebők. After evaluation in the nursery and orchard tests there are four promising rootstock cultivars selected from this material: 'Korponay' used as self fertile seed tree, its seedlings are recommended for sour cherries. The mahaleb varieties 'Bogdány' (vigorous), 'Egervár' and 'Magyar' (medium vigour) are propagated by cuttings. The next project has started in 1979 with the aim to select self fertile mahaleb seed trees producing homogeneous seedling populations with reduced vigour. Inbred populations from isolated flowering self fertile trees were produced and planted out in 1980. The inbreeding of 'Korponay' self fertile P. mahaleb variety resulted in specimens with different fruit colour (yellow, red, black), fruit shape and size. From among them self fertile trees were selected with various growth characteristics. Seedlings of that self fertile mother trees (S2 population) were tested in seedbed, they showed homogeneous phenotype characteristics as liners in the nursery. As rootstock of 'Érdi bőtermő' sour cherry in the orchard most of the S2 lines proved to be less vigorous in comparison to SI popuplations. 'Érdi bőtermő' trees budded on certain S2 lines in the orchard are more productive than those on S1 ('Korponay' seedling). Characteristics of the S2 generation as seed tree were studied as well. We expect to get morphologically homogeneous seedling populations with different growth vigour and good productivity in the later inbred generations. In the last couple of decades the research activity concerning ground-cherry and its hybrids resulted in dwarfing rootstocks. Prunus fruticosa Pall. hybrids from the natural flora of Hungary were collected and artificial hybrids were created between P. fruticosa and mahaleb cherry. Most of them are in the initial tests, only one of them is before registration, named 'Prob', which is a dwarf rootstock for sweet cherry. By the screening of new hybrids medium vigorous or semi dwarfing and precocious rootstocks seem to be promising for the cherry industry.
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Concentration of ‘Oblachinska’ sour cherry juice using osmotic distillation
31-34.Views:229Sour cherry is one of the most famous and characteristic Hungarian fruit what is also commonly used in food production as a colour additive. The sour cherry cultivar, ‘Oblachinska’ was shown to be a rich source of antioxidant compounds, including mainly anthocyanins. Valuable compounds in fruit juice – vitamins, polyphyenols etc. –are heat-sensitive molecules, which should be taken into account during the process of concentration to prevent degradation. Osmotic distillation seems a suitable option to product high quality sour cherry juice because this process does not require high temperature or pressure. Raw juice with approximately 15°Brix was used for the experiment and tried to concentrate up to 60° Brix where the water activity low enough to inhibit the microbiological deterioration. Before and after the process, TPC (total phenolic compounds) and FRAP (ferric reducing antioxidant power) was measured using spectrophotometric methods to determine the effect of the osmotic distillation. Our results point out that osmotic distillation is a promising method to concentrate sour cherry juice and prevent the loss of valuable compounds.
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An investigation of genotype-environment interaction and stability for pea (Pisum sativum L) seed yield
53-55.Views:107Eighteen pea breeding lines and three check cultivars were tested in two years with and without irrigation. The linear regression can explain only a small part of GEI so the use of regression technique was not possible for interpreting the data. In this case the ecovalence, stability variance and superiority measure stability parameters cannot describe properly the genotype's response. With the AMMI method it is possible to group properly the genotypes according to their response.
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Effect of different cultivation methods on strawberry’s antioxidant value
7-10.Views:374The small fruit and strawberry production is economically important in Serbian agriculture. The fresh fruit is sold in domestic markets, but the frozen and processed is mainly sold in foreign markets. The growers find themselves in an ever-expanding selection of the varieties, but besides the nursery and breeder’s descriptions of new cultivars, there are only few exact results for the biological and economical ability of the varieties, which makes easier to choose the proper cultivar. The performance of the varieties in modern strawberry production is worth to investigate by adaptive cultivation methods and production technique. In the case of new strawberry varieties we know neither their needs in production nor their real capability, so they have to be compared with an older, standard control variety (‘Clery’). The breeder’s manuals give only references for the needs of the production technique, of the variety, so it is necessary to have facts, which are based on objective testing results. In this work, a new variety (‘Joly’) is being compared with the control variety. Comparative study of individual varieties are required covering the antioxidant content of the fruits (FRAP) and variability of these amounts in different years of breeding as well as the production techniques under similar circumstances.
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Micropropagation of Rudbeckia hirta L. from seedling explants
105-108Views:162We conducted experiments for developing an in vitro micropropagation protocol starting from meristems of Rudbeckia hirta L seedlings. We pre-soaked the seeds in sterile ion-exchanged water for 17 hours, and then achieved surface disinfection in two separate steps. First, we used concentrated household sodium-hypochloride solution for 20 minutes and, also for 20 minutes, we applied hydrogen peroxide of 10%, which was followed by washing with sterile ion-exchanged water three times. For the propagation of seedling meristems, the combination of half-strenght solid Murashige and Skoog (1962) culture medium containing 10 mg/1 of kinetin or 2 mg/I of kinetin + 0.1 mg/1 of 2iP proved to be the most suitable. The average number of shoot-buds developed from the seedling axillary meristem in the best culture media varied between 5 and 17. Without separating them, we inoculated the shoot-bud clusters on MS culture medium containing 2 mg/1 of IAA. After four weeks of incubation we obtained elongated shoots which we separated and inoculated into a new culture medium and we obtained elongated roots. The rooted plants were gradually acclimatised in the cultivation room, potted and carried to a greenhouse, and then planted in open field for subsequent observation. By adopting this method, our laboratory started the micropropagation of the superior and/or elite genotypes of the Rudbeckia hirta L. being of special value in respect of breeding.