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The aim of this diploma thesis was to evaluate selected parameters indicating the quality of dried pea seeds (Pisum sativum L.) in selected group of two varieties. Literary research focuses mainly on pea production in the Czech Republic and worldwide, for breeding varieties, peas and seed recognition. The literary research directly follows the practical part, which deals with the process of recognition and certification of seeds of two varieties in practice. In addition, laboratory tests of pea seed quality, especially germination. This was set as part of the sub-supply, supply and cleanse seeds, for the Gambit variety it recorded 69% and fell to 37% for the Salamanca variety, which differed considerably from the pre-supply and delivery seeds. This research shows an internal view within the farm and the great complexity and difficulty of the seed certification process.

Pterocarpan phytoalexins are antimicrobial compounds produced by legumes when challenged by biotic stresses. Most legumes produce pterocarpan phytoalexins with (-)-stereochemistry but pea (Pisum sativum L.) produces as its major phytoalexin (+)-pisatin. Pea also occasionally produces a minor amount of (-)-maackiain as a pterocarpan phytoalexin, and studies on the biosyntheses of (+)-pisatin and (-)-maackiain have shown that up to (-)-7,2'-dihydroxy-4',5'-methylenedioxyisoflavanone [(-)-sophorol] and 7,2'-dihydroxy-4',5'-methyl-enedioxyisoflavanol [(-)-DMDI]they have common intermediates with (-)-DMDI being where the two pathways diverge. The final step in (+)-pisatin biosynthesis is the methylation of (+)-6a-hydroxymaackiain [(+)-6a-HMK] by 6a-hydroxymaackiain methyltransferase (HMM2) but the steps from (-)-DMDI to (+)-6a-HMK are unknown.The shifting of the stereochemistry from (-)-DMDI to (+)-6a-HMK has been proposed to involve the achiral isoflavene, 7, 2'-dihydroxy-4', 5'-methylene-dioxyisoflav-3-ene (DMDIF). In this dissertation, I have shown that cis-(-)-DMDI is the enzymatic product of (-)-sophorol, and is the precursor of DMDIF which is produced by the dehydration activity of "isoflavene synthase" (IFVS). IFVS activity was not observed in elicited tissues of alfalfa, chickpea, beans, pepper, and broccoli, plants that do not produce (+) pterocarpans. Partial purification of IFVS demonstrated that it is either large in size or tightly complexed with other proteins. The SDS-PAGE of the 29-fold purified product revealed 12 major bands that aggregated into 3 bands in the non-denaturing PAGE. IFVS activity was in band 3 which co-migrated with marker proteins of>100 kDa in size. Proteins identified from LC-MS/MS peptide sequences of the proteins in band 3, when compared to three protein databases, did not identify any proteins with an enzymatic activity expected for IFVS. A disease resistance-response protein (a dirigent-like protein) and two protein-binding proteins were the most abundantly detected proteins in the pea transcriptome-translated database. Also, four of the known enzymes (isoflavone reductase, HMM1, HMM2, and sophorol reductase) involved in (+)-pisatin biosynthesis were among the proteins identified. It may be that IFVS is associated with these other proteins as a complex in vitro and in vivo. The lack of detection of IFVS in the databases could be because it has not yet been sequenced as it functions in a rare biosynthetic pathway.

The aim of this thesis is the study of the genetic substance of the important domestication sign pod dehiscence. Two types of pea were analyzed, with indehiscence pods JI92 (Pisum sativum subsp. sativum) and wild field pea with dehiscent pods JI64. (Pisum sativum subsp. elatius). By reciprocal crossbreeding of these two lines, were created recombinant inbred lines (RILs), of a total of 134 RILs lines were selected with 9 contrast lines. We utilized the massive parallel sequencing of the 3'ends of the cDNA, obtained by reverse transcription of mRNA isolated from the seam. Thanks to this method, 3 candidate genes were generated. Subsequently, we determined the expression of these three candidate genes for the using quantitative Real-Time PCR (RT-qPCR). Amplification curves and Ct values generated from the RT-qPCR were subsequently used to generate graphs to show the degree of expression of the candidate genes. The most suitable candidate was the Ps15 gene, which is present in LGIII in the Dpo1 region, and therefore could be responsible for pod dehiscence.

Made available in DSpace on 2014-08-20T13:59:08Z (GMT). No. of bitstreams: 1 dissertacao_marcio_espinosa_de_farias.pdf: 3645960 bytes, checksum: 2a09be33a4c426dea1aef53c4073f253 (MD5) Previous issue date: 2014-02-17 / Photosynthesis plays a fundamental role in the processes of growth and development of plants. It occurs in the chloroplasts of leaf mesophyll cells and consists of two steps, a photochemical phase that converts light energy into chemical energy, and other biochemical phase, which uses the energy of the first step for fixing CO2. The study aimed to evaluate, in detached leaves of pea (Pisum sativum L.), the behavior of the electron transport chain in the presence of inhibitors of specific points. The plants were grown in a greenhouse. Two to four weeks after sowing, when the leaves had become fully expanded, they were detached and the petioles were immersed in solutions containing 0, 25, 50, 100, 250, and 500 μM of DCMU, atrazine, DBMIB and methyl viologen. To DCMU, atrazine and methyl viologen the leaves remained for 2 h in contact with the solutions, to DBMIB was 5h. Measurements of transient fluorescence of chlorophyll, delayed fluorescence and modulated reflection at 820nm were made using M-PEA fluorometer. The leaves were dark adapted for 30 minutes, before a saturating pulse emission, and the fluorescence intensities were measured for 60 seconds. Pea leaves treated with 500 μM concentration of DCMU and atrazine showed to be sensitive to this dose of inhibitors, in analyzing the results of transient fluorescence, delayed fluorescence and modulated reflection at 820 nm. The leaves treated with 500 μM of methyl viologen showed differences in parameters related to the mechanism of action of the inhibitor.The same concentration used to DBMIB, had no significant difference in many results compared to the control, only in those involved in the reduction of end acceptors of photosystem I. Therefore, a detailed analysis of the transient fluorescence, delayed fluorescence and modulated reflection at 820nm, allows us to collect and correlate a series of information about the whole electron transport chain. / A fotossíntese exerce um papel fundamental para os processos de crescimento e desenvolvimento das plantas. Ela ocorre nos cloroplastos das células do mesofilo foliar e consiste de duas etapas, uma fase fotoquímica, que converte energia luminosa em energia química e outra fase bioquímica, que utiliza a energia da primeira etapa pra a fixação do CO2. O trabalho teve como objetivo avaliar, em folhas destacadas de ervilha (Pisum sativum L.), o comportamento da cadeia de transporte de elétrons na presença de inibidores de pontos específicos. As plantas foram cultivadas em casa de vegetação. Duas a quatro semanas após semeadura, quando as folhas apresentavam-se completamente expandidas, elas foram destacadas e os pecíolos foram imersos em soluções contendo 0, 25, 50, 100, 250 e 500 μM de DCMU, atrazina, DBMIB e metil viologênio. Para DCMU, atrazina e metil viologênio as folhas permaneceram por 2h em contato com as soluções, para o DBMIB foi de 5h. As medidas da fluorescência transiente da clorofila, da decaída de fluorescência e da reflexão modulada à 820nm, foram feitas utilizando fluorômetro M-PEA. As folhas foram adaptadas ao escuro por 30 minutos, antes da emissão de um pulso saturante, e as intensidades de fluorescência foram medidas por 60 segundos. As folhas de ervilha tratadas com a concentração de 500 μM de DCMU e atrazina demonstraram ser sensíveis a essa dose dos inibidores, ao analisar os resultados da fluorescência transiente, decaída de fluorescência e da reflexão modulada a 820 nm. As folhas tratadas com 500 μM de metil viologênio apresentaram diferenças nos parâmetros relacionados com o mecanismo de ação do inibidor. A mesma concentração utilizada para DBMIB, não teve diferença em muitos resultados em relação ao controle, apenas naqueles envolvidos com a redução dos aceptores finais do fotossistema I. Dessa maneira, uma análise detalhada da fluorescência transiente, da decaída da fluorescência e da reflexão modulada a 820 nm, permite coletar e correlacionar uma série de informações sobre toda a cadeia de transporte de elétrons.

La connaissance des effets et modes d’action des QTL (Quantitative Trait Loci) de résistance des plantes aux maladies est un enjeu majeur pour l’exploitation durable des résistances génétiques en agriculture. Dans le pathosystème pois/Aphanomyces euteiches, présentant une importance économique majeure, l’objectif de la thèse a consisté à valider les effets de QTL de résistance, seuls ou en combinaison, et à connaitre leurs modes d’action sur les étapes du cycle de l’agent pathogène. Un total de 157 NILs (Near Isogenic Lines) de pois issues d’un programme de Back-cross Assisté par Marqueurs, porteuses d’aucun, un, deux ou trois des sept principaux QTL de résistance préalablement identifiés, a été génotypé à l’aide d’une puce 15K SNPs et évalué pour la résistance. Les NILs porteuses de QTL à effets majeurs, seuls ou en combinaison avec des QTL à effets mineurs,ont présenté des niveaux accrus de résistance partielle en conditions contrôlées et au champ par rapport aux NILs dépourvues de QTL, dans différents fonds génétiques. Certaines NILs comportant des QTL individuels ou combinés à effets mineurs ont également montré un niveau réduit de sévérité de la maladie dans l’une ou les deux conditions de test. La plupart des QTL a présenté des effets significatifs sur le ralentissement de l’infection et/ou de la quantité d’ADN pathogène ayant colonisé la racine pendant sept jours après inoculation. Cette thèse fournit des outils et éléments de choix de QTL à combiner en sélection pour augmenter l’efficacité de la résistance partielle à A. euteiches dans les futures variétés de pois / Knowledge of the effects and action modes of resistance QTL to plant diseases is a major challenge for the durable use of genetic resistances in agriculture. In the pea/Aphanomyces euteiches pathosystem, which has a major economic importance, the aim of this study was to validate the single or combined effects of main resistance QTL, and study their action modes on steps of the pathogen life cycle. A total of 157 pea NILs (Near Isogenic Lines) created by Marker-Assisted Back-crossing and carrying no, one, two or three of the seven main resistance QTL previously identified, was genotyped using a 15K SNPs array and evaluated for resistance. The NILs carrying major-effect QTL, individually or in combination with minor-effect QTL, had increased levels of partial resistance in controlledconditions and in the field compared to NILs without QTL, in different genetic backgrounds. Several NILs carrying single or multiple minor-effect QTL also showed reduced levels of disease severity in one or the two test conditions. Most of the QTL had significant effects on slowing down infection and/or pathogen DNA quantity which had colonized the root for seven days after inoculation. This study gives tools and information for the choice of resistance QTL to use in pyramiding breeding strategies for increasing partial levels of resistance to A. euteiches in future pea varieties.

The diploma thesis deals with the determination of mobile forms of mercury using the technology of diffusion gradient in thin-film. The theoretical part deals with the occurrence of chemical forms of mercury in the environment, mercury effects on plants and methods of determination of mercury's mobile forms. In the practical part, the accumulation of mercury from soils into various organs of garden pea (Pisum sativum L.) was observed. Soil samples were taken in Brno, in streets Opuštěná and Zvonařka. The contents of mercury in soils ranged from 0,32618 +- 0,02493 mg.kg-1 to 0,08382 +- 0,00255 mg.kg-1. Only 0,15 - 0,20 % of the mercury passed from the soil into the soil solution. Only 2,21 to 3,45 % of mercury was available for the soil solution for DGT units and therefore also for the plants. The mercury content in the soil influenced the amount of mercury in the garden pea. The highest concentrations of mercury were detected in the leaves and roots, the lowest concentrations were in the consumed parts of garden pea. The significant correlations were found out between the content of mercury's mobile forms provided by DGT method and the mercury content in roots, leaves and stem of garden pea (correlation coefficients from 0,913 to 0,984). Simulation of acid rains did not demonstrate the raise of mercury's mobility relating to the increase of the acidification of the soil.

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