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  • About
  • The Global ETD Search service is a free service for researchers to find electronic theses and dissertations. This service is provided by the Networked Digital Library of Theses and Dissertations.
    Our metadata is collected from universities around the world. If you manage a university/consortium/country archive and want to be added, details can be found on the NDLTD website.
1

Analysis of inflammatory changes in human pancreatic islet cells

Jackson, Andrew M. Naziruddin, Bashoo. January 2009 (has links)
Thesis (Ph.D.)--Baylor University, 2009. / Includes bibliographical references (p. 126-143).
2

Rétroactions positives et mémoire cellulaire : exemples dans l'expression génétique et le métabolisme cellulaire / Positive feedback loops and cellular memory : examples in gene expression and cellular metabolism

Nicol-Benoit, Floriane 03 July 2013 (has links)
Au-delà de l'information génétique contenue dans la séquence de l'ADN des cellules, il existe une mémoire cellulaire, dite épigénétique comprenant l'ensemble des circuits génétiques avec rétroactions positives permettant d'amplifier ou de maintenir une réponse cellulaire dans le temps. Nous nous sommes intéressés, à travers deux exemples, aux boucles de rétrocontrôle positif comme élément de réponse à un signal, permettant de fixer, de manière à la fois dynamique et robuste, le comportement cellulaire. Dans un premier temps, nous avons identifié une boucle d'auto-amplification dans la production de vitellogénine chez la truite et permettant d'expliquer l' « effet mémoire de la vitellogénèse » (une seconde stimulation à l'œstradiol induit une plus forte production de vitellogénine et plus rapidement que lors de la première stimulation, alors même que le niveau de vitellogénine retombe à zéro entre les deux stimulations). Le modèle que nous proposons implique un récepteur tronqué à l'œstradiol possédant une activité basale même en l'absence de son ligand, permettant de maintenir la cellule dans un état d'aptitude à répondre sans pour autant produire de vitellogénine. Dans un deuxième temps, nous nous sommes intéressés à une des causes possibles provoquant la transition épithélio-mésenchymateuse (EMT), responsable des métastases dans les cancers. L'EMT témoigne d'un état plus agressif des cellules tumorales et s'accompagne notamment d'un changement du métabolisme des cellules cancéreuses, diminuant la part de la phosphorylation oxydative au profit de la glycolyse (effet Warburg). Cela entraîne une baisse d'efficacité de la production d'ATP, obligeant les cellules à prélever davantage de nutriments dans leur milieu. Cette observation a suscité le développement de thérapies basées sur la privation de glucose et qui, a priori, devraient nuire principalement aux cellules cancéreuses. Nous avons étudié les effets d'un faible contenu cellulaire en ATP sur la transformation cellulaire. Nous avons observé qu'un traitement par un analogue non métabolisable du glucose diminue drastiquement le contenu en ATP des cellules ayant passé l'EMT et induit des changements morphologiques et génétiques orientés vers le phénotype mésenchymateux. La protéine MKL1, cofacteur de transcription dont l'activité est régulée par la polymérisation de l'actine, pourrait être un relais génétique entre l'état métabolique cellulaire et le maintien de l'EMT. Ces résultats suggèrent de fortes connections entre l'EMT et le niveau énergétique des cellules, faisant d'une privation d'énergie une cause possible de l'aggravation du phénotype mésenchymateux et remettant en cause les bienfaits sur le long terme de thérapies visant à « affamer » les cellules tumorales. / Beyond the genetic information contained in the DNA sequence of cells, there is a cellular memory called epigenetic, including genetic circuits with positive feedback loops amplifying or maintaining cellular states in time. We studied through two examples, the positive feedback loops as part of response to a signal, able to set cell behavior, in a dynamic and robust way. As a first step, we identified a self-amplification loop in the production of trout vitellogenin explaining the "vitellogenesis memory effect" (a second estradiol stimulation induces higher and faster vitellogenin production than during the first stimulation, even though the vitellogenin level falls to zero between the two stimuli). The model we propose involves a truncated estradiol receptor, with a basal activity even in the absence of its ligand, which is able to maintain the cell in an estrogen-responsive state without producing vitellogenin. In a second step, we studied one of the possible causes leading to the epithelial-mesenchymal transition (EMT), involved in cancer metastasis. The EMT reflects a more aggressive state of tumor cells and is associated with a particular change in the metabolism of cancer cells, reducing the part of oxidative phosphorylation in favor of glycolysis (Warburg effect). This leads to a reduction in the efficiency of ATP production, forcing the cells to take more nutrients from their environment. This observation led to the development of treatments based on glucose deprivation which should mainly affect cancer cells. We studied the effects of a low cellular ATP content on cell transformation. We observed that a treatment with a non-metabolizable glucose analogue drastically reduces the ATP content of cells that had undergone EMT and induces morphological and genetic changes enforcing the mesenchymal phenotype. We identified the transcriptional coactivator MKL1, whose activity is regulated by actin polymerization, as a possible genetic link between the cellular metabolic state and maintenance of EMT. These results suggest strong connections between the EMT and the energy level of the cells, and raise serious questions about the benefits of the long-term therapy "starving" tumor cells, considering that energy deprivation could aggravate the mesenchymal cell phenotype.
3

PREDICTION OF THE NGA-WEST2 AVERAGE HORIZONTAL PEAK GROUND ACCELERATION USING GENETIC EXPRESSION PROGRAMMING

Sanad, Abdel-Aziz 01 May 2022 (has links)
Genetic Expression Programming (GEP) is used to create Ground Motion Predicting Equations (GMPEs) for the average peak ground acceleration using 12,854 ground motion records obtained from the NGA-WEST2 project. The predictor set considered in this research consists of the moment magnitude, dip angle, rake angle, depth to the top of fault rupture, Joyner Boore distance, closest distance to the ruptured fault area, and the shear wave velocity in the top 30 m of the site. Four out of 23 candidate models were able to fairly predict the PGA for magnitudes larger than 4.5 and compared well with existing GMPEs in literature. GEP was capable of reasonably predicting the physical importance of the magnitude and distance parameters. However, other parameters often were either not fitted, or fitted as regression coefficients. The results illustrate GEP’s potential as a viable alternative to regression methods currently used in developing GMPEs.
4

Discovering unknown equations that describe large data sets using genetic programming techniques

González, David Muñoz January 2005 (has links)
<p>FIR filters are widely used nowadays, with applications from MP3 players, Hi-Fi systems, digital TVs, etc. to communication systems like wireless communication. They are implemented in DSPs and there are several trade-offs that make important to have an exact as possible estimation of the required filter order. </p><p>In order to find a better estimation of the filter order than the existing ones, genetic expression programming (GEP) is used. GEP is a Genetic Algorithm that can be used in function finding. It is implemented in a commercial application which, after the appropriate input file and settings have been provided, performs the evolution of the individuals in the input file so that a good solution is found. The thesis is the first one in this new research line. </p><p>The aim has been not only reaching the desired estimation but also pave the way for further investigations.</p>
5

Discovering unknown equations that describe large data sets using genetic programming techniques

González, David Muñoz January 2005 (has links)
FIR filters are widely used nowadays, with applications from MP3 players, Hi-Fi systems, digital TVs, etc. to communication systems like wireless communication. They are implemented in DSPs and there are several trade-offs that make important to have an exact as possible estimation of the required filter order. In order to find a better estimation of the filter order than the existing ones, genetic expression programming (GEP) is used. GEP is a Genetic Algorithm that can be used in function finding. It is implemented in a commercial application which, after the appropriate input file and settings have been provided, performs the evolution of the individuals in the input file so that a good solution is found. The thesis is the first one in this new research line. The aim has been not only reaching the desired estimation but also pave the way for further investigations.
6

Efeito de um agonista dos receptores ativados por proliferadores de peroxissomo gama (PPARγ) sobre os efeitos do ácido linoleico conjugado (CLA, trans-10, Cis-12 e cis-9, trans-11) na transcrição de genes lipogênicos em explantes mamários de ovelhas lactantes / Effect of as activated agonist receptor by proliferators of gama peroxisome about the effects of linoleic acid on lipogenic genes in mammary explants of lactaing sleeps

Brogin Junior, Wagner 22 February 2017 (has links)
Submitted by Claudia Rocha (claudia.rocha@udesc.br) on 2018-03-16T13:45:25Z No. of bitstreams: 1 PGCA17MA219.pdf: 853811 bytes, checksum: 5ed849be56df67865d9463e92b006618 (MD5) / Made available in DSpace on 2018-03-16T13:45:25Z (GMT). No. of bitstreams: 1 PGCA17MA219.pdf: 853811 bytes, checksum: 5ed849be56df67865d9463e92b006618 (MD5) Previous issue date: 2017-02-22 / Capes / The search for optimization of production systems with animals leads to besides the animal fator, and show us that the micro components are the ones which sustain the production factores. On this way the nutrigenomic animal gains more importance and prominent in the search forresourse efficiency. Thereby the aim of this work was to analyse the effects of peroxisome proliferator activated gamma receptors (PPARγ) in the transcription of lipogenic genes and its response to conjugated linoleic acid (mixture of isomers trans-10, cis12 and cis-9, trans-11) through a specific chemical agonist (TZD, thiazolinedione). 1) Control: growing medium, 400μl; 2) TZD: 40ml (10μMol/Lt); 3) CLA (50% of CLA trans10, cis-12 and 50% of CLA cis-9, trans-11): 30ml (315μMol/Lt) and 4) TZD+CLA: 40ml TZD (10μMol/Lt) + 30ml (315μMol/Lt). It was extracted the RNA, synthesized the complementary DNA (cDNA), and carried out the polimerase chain reaction in Real Time (PCR – Real Time), measured the genetic expression of isoform PIII of acetyl-CoA-carboxylase alpha (ACCα), fatty acid synthase (FASN), peroxisome proliferator activated gamma receptors (PPARγ), sterol regulatory elemento-binding protein 1 (SREBP1), cleavage activation protein of SREBP1 (SCAP), stearoyl-CoA-desaturase (SCD), insulin induced gene 1(INSIG1), insulin induced gene 2(INSIG2). Compared to control, the treatment TZD increased the genes expression being SREBP1(1010%), INSIG1 (789%), INSIG2 (849%), FASN (7831%), ACCα (8753%), SCD (6272%) and PPARγ (620%). Compared to the TZD+CLA treatment, the TZD treatment increased the expression of the genes SREBP1 (237%), ACCα (1729%), INSIG1 (7142%) and PPARγ (2480%). Compared to CLA treatment, TZD increased the expression of FASN (275%), SCAP (916%), SCD (206%) and INSIG2 (1700%). It was concluded that the use of chemical agonist TZD in explants of the mammary gland of ewes, grown in vitro has acted to induce a greater expression of PPARγ and CLA reduces the genes expression FASN, ACCα, SCD and SCAP. In the TZD+CLA treatment, the TZD does not exceed the capacity of reducing CLA gene expression for the PPARγ, FASN, SCD, SCAP, INSIG1 and INSIG2 / A busca pela otimização dos sistemas produtivos com animais conduz para além do fator animal, e nos mostram que os componentes “micro” são que sustentam os fatores de produção. Deste modo a nutrigenômica animal ganha cada vez mais importância e destaque na busca pela eficiência de recursos. Com isso, o presente trabalho visou analisar o efeito dos receptores ativados por proliferadores de peroxissomo gama (PPARγ) na transcrição de genes lipogênicos e sua resposta ao ácido linoleico conjugado (CLA, com os isômeros trans-10, cis-12 e cis-9, trans-11), através de um agonista químico específico (TZD, thiazolinediona). No experimento foram realizados cultivos de explantes de glândula mamária de ovelhas lactantes, submetidos aos seguintes tratamentos: 1) Controle: meio de cultivo, 400μl; 2) TZD: 40mL (10 μMol/Lt); 3) CLA: (50% de trans-10, cis-12 e 50% de cis-9, trans-11): 30mL (315 μMol/Lt) e; 4) TZD + CLA: 40mL TZD (10 μMol/Lt) + 30mL (315 μMol/Lt). Foi extraído o RNA, sintetizado o DNA complementar (cDNA) e realizado a reação da cadeia da polimerase em tempo real (PCR Real Time) e medida a expressão gênica da isoforma PIII da acetil-CoA-carboxilase alfa (ACCα), ácido graxo sintase (FASN), receptores ativados por proliferadores de peroxissomo gama (PPARγ), proteína de ligação ao elemento de resposta a esterol 1 (SREBP1), proteína de ativação de clivagem da SREBP1 (SCAP), estearoil-CoA-dessaturase (SCD), gene-1 induzido pela insulina (INSIG1), gene-2 induzido pela insulina (INSIG2). Comparado ao Controle, o TZD aumentou a expressão dos genes, sendo SREBP1 (1.010%), INSIG1 (789%), INSIG2 (849%), FASN (7831%), ACCα (8753%), SCD (6272%) e PPARγ (620%). Comparado ao tratamento TZD+CLA o TZD, aumentou a expressão dos genes SREBP1 (237%), ACCα (1729%), INSIG1 (7142%) e PPARγ (2480%). Em comparação ao tratamento CLA, o TZD aumentou a expressão da FASN (275%), SCAP (916%), SCD (206%) e INSIG2 (1700%). Conclui-se que o uso do agonista químico TZD em explantes da glândula mamária de ovelhas lactantes aumenta a expressão do PPARγ e o CLA reduz a expressão gênica da FASN, ACCα, SCD e SCAP. No tratamento TZD+CLA, o TZD não supera a capacidade de redução da expressão gênica do CLA para os genes PPARγ, FASN, SCD, SCAP, INSIG1 e INSIG2
7

Expressão das proteínas N e P do vírus respiratório sincicial humano: estudos funcionais e de imunização. / Expression of human respiratory syncytial virus N and P proteins: functional and immunization studies.

Simabuco, Fernando Moreira 12 February 2009 (has links)
O Vírus Respiratório Sincicial Humano é um vírus envelopado de RNA negativo e considerado o patógeno mais importante do trato respiratório de bebês e crianças. As proteínas virais N e P foram expressas em bactérias, purificadas e usadas para a produção de anticorpos policlonais em camundongos. Estudos in silico permitiram a predição de regiões desordenadas da proteína P, as quais foram identificadas por espectrometria de massa como regiões hiper sensíveis a proteases. A otimização dos genes N e P permitiu uma forte e eficiente expressão das proteínas N e P em células humanas. Vacinas de DNA contendo os genes otimizados foram testadas em camundongos e geraram forte resposta imune humoral. As proteínas N e P expressas em células humanas foram imunoprecipitadas e analisadas quanto a interações com proteínas celulares, identificadas por espectrometria de massa. A proteína P mostrou ser capaz de interagir com a HSP70. Por fim, um sistema Minigenoma alternativo, usando o promotor da RNA polimerase II, foi construído para o HRSV mas pouca ou nula atividade foi detectada. / The Human Respiratory Syncytial Virus is a single stranded negative RNA enveloped virus and it is considered the most important pathogen of the respiratory tract of infants and neonates. The viral N and P proteins were expressed in bacteria, purified and used for the production of polyclonal antibodies in mice. In silico studies allowed the prediction of intrinsically disordered domains for P protein, which were identified by mass spectrometry as protease hyper sensible regions. The optimization of N and P genes allowed a robust expression of the N and P proteins in human cells. DNA vaccines containing the optimized genes were tested in mice and generated strong humoral imune response. The N and P proteins expressed in human cells were immunoprecipitated and their interactions with cellular proteins were identified by mass spectrometry. The P protein was able to interact with the HSP70 protein. Finally, an alternative Minigenome system, using an RNA polymerase II promoter, was developed for HRSV but low or no activity was detected.
8

Análise estrutural e funcional da região promotora de rDNA de Leishmania (Viannia) braziliensis e estudo comparativo com as regiões de Leishmania (Leishmania) / Structural and functional caracterization of rDNA promoter region of Leishmania (Viannia) braziliensis and comparative study with the Leishmania (Leishmania) region

Nassar, Maira Natali 08 May 2009 (has links)
Um conjunto de quadros clínicos conhecidos genericamente por Leishmaniose, que representa um sério problema de Saúde Pública, tem como agentes etiológicos protozoários do gênero Leishmania. Em seu ciclo de vida, o parasita apresenta dois hospedeiros, um vertebrado e um invertebrado. O gênero Leishmania é dividido em dois subgêneros: Leishmania (Leishmania) e LeishmaniaViannia), de acordo com a posição ocupada pela forma promastigota no tubo digestivo do hospedeiro invertebrado. Genes que codificam RNA ribossômico são exclusivamente transcritos pela RNA polimerase I. A região promotora dessa polimerase é conhecida como espécie-específica. Estudos anteriores, baseados em ensaios de expressão transiente com construções nas quais a expressão do gene repórter CAT era dirigido por diferentes regiões do promotor mostraram que em Leishmania há um reconhecimento espécie-específico, mas que espécies filogeneticamente relacionadas reconhecem melhor o promotor do que a espécie homóloga. A caracterização estrutural da região promotora de RNA pol I de L. (V.) braziliensis possibilitou mapear o sítio de início de transcrição e definir a provável região promotora de RNA pol I desse organismo. Quando comparamos o 5´ ETS da região estudada, com a região correspondente das demais espécies do subgênero L. (Leishmania), notamos uma alta similaridade tanto no tamanho, quanto na seqüência de nucleotídeos. Já na região IGS os blocos de repetição apresentam tamanho similar, porém seqüências distintas, assim como a seqüência à montante do ETS. Não foi determinado o número de blocos de repetição presente em cada cístron de L.(V.) braziliensis. Os estudos de ligação de fatores nucleares à região central do promotor mostraram que houve um reconhecimento diferencial dessa região entre a espécie homóloga L. (V.) braziliensis e as espécies heterólogas L. (V.) guyanensis, pertencente ao mesmo subgênero e L. (L.) amazonenis. Os complexos protéicos associados a essa região central apresentaram maior afinidade com a espécie heteróloga L. (V.) guyanensis. O fato de ser o ensaio de CAT trabalhoso e demorado levou à busca de outro repórter que evidenciasse a funcionalidade da região promotora ao mesmo tempo em que permitisse a quantificação dessa expressão, de modo a medir a força do promotor. Neste trabalho iniciamos a padronização para utilizar GFP como gene repórter no estudo da funcionalidade de promotores. Foi possível mostrar que a GFP é detectável em microscopia confocal e que as células que expressam GFP podem ser quantificadas em FACS, no entanto, essas detecções só foram possíveis em parasitas selecionados por uma marca de seleção, ou seja, numa transfecção estável. Assim, substituir o gene repórter CAT pelo gene GFP foi inadequado para os ensaios de transfecção transiente, impedindo que fosse medida a atividade da região promotora de L. (V.) braziliensis em diferentes espécies. / Leishmaniasis is the generic name of a serious public health problem that is caused by protozoa of the genus Leishmania. In its lifecycle, the parasite has two hosts, a vertebrate and an invertebrate. The genus Leishmania is divided into two subgenera: Leishmania (Leishmania) and Leishmania (Vianna), according to the position occupied by the promastigotes form at the gut of the invertebrate host. Genes that encode ribosomal RNA are exclusively transcribed by RNA polymerase I. The promoter region of the RNA polymerase I is known to present a species-specific recognition. Previous studies, based on transient expression assays with constructions in which the expression of the reporter gene CAT was directed by different regions of the promoter showed that RNA pol I promoter in Leishmania is species-specific, but in phylogenetically related species the expression of the reporter gene is higher than in the homologous species. The structural characterization of the promoter region of RNA pol I of L. (V.) braziliensis enabled to map the site of initiation of transcription and to define the promoter region of RNA pol I. The comparison of the determined 5\' region of the ETS region, with the corresponding region of other species of the subgenus L. (Leishmania), showed a high similarity both in size, as in the sequence of nucleotides. However, the IGS region and the repetitive blocks of nucleotides have similar size but different sequences. The studies of binding of nuclear factors to the central region of the promoter showed that there was a recognition between the species counterpart L. (V.) braziliensis and the heterologous species L. (V.) guyanensis, belonging to the same subgenus and L. (L.) amazonensis, that belongs to another subgenus. The complex protein associated with this central region showed greater affinity with the heterologous species L. (V.) guyanensis. The fact that to quantify the CAT expression represents a laborious and time consuming test lead us to search for another reporter that could identified the functionality of the promoter region and at the same time allowed the quantification of expression in order to measure the strength of the promoter. In the present work, we began the standardization for the use of GFP gene as reporter in the study of the functionality of promoters. It was possible to show that GFP is detectable in confocal microscope and the cells that express GFP can be quantified in FACS, however, these findings were made possible only in parasites selected by the mark, ie in a stable transfection. So, the replacement of CAT by GFP reporter showed to be inadequate for testing promoters in transient transfection. This prevented the measure of the activity of the promoter region of L. (V.) braziliensis in different species.
9

Análise estrutural e funcional da região promotora de rDNA de Leishmania (Viannia) braziliensis e estudo comparativo com as regiões de Leishmania (Leishmania) / Structural and functional caracterization of rDNA promoter region of Leishmania (Viannia) braziliensis and comparative study with the Leishmania (Leishmania) region

Maira Natali Nassar 08 May 2009 (has links)
Um conjunto de quadros clínicos conhecidos genericamente por Leishmaniose, que representa um sério problema de Saúde Pública, tem como agentes etiológicos protozoários do gênero Leishmania. Em seu ciclo de vida, o parasita apresenta dois hospedeiros, um vertebrado e um invertebrado. O gênero Leishmania é dividido em dois subgêneros: Leishmania (Leishmania) e LeishmaniaViannia), de acordo com a posição ocupada pela forma promastigota no tubo digestivo do hospedeiro invertebrado. Genes que codificam RNA ribossômico são exclusivamente transcritos pela RNA polimerase I. A região promotora dessa polimerase é conhecida como espécie-específica. Estudos anteriores, baseados em ensaios de expressão transiente com construções nas quais a expressão do gene repórter CAT era dirigido por diferentes regiões do promotor mostraram que em Leishmania há um reconhecimento espécie-específico, mas que espécies filogeneticamente relacionadas reconhecem melhor o promotor do que a espécie homóloga. A caracterização estrutural da região promotora de RNA pol I de L. (V.) braziliensis possibilitou mapear o sítio de início de transcrição e definir a provável região promotora de RNA pol I desse organismo. Quando comparamos o 5´ ETS da região estudada, com a região correspondente das demais espécies do subgênero L. (Leishmania), notamos uma alta similaridade tanto no tamanho, quanto na seqüência de nucleotídeos. Já na região IGS os blocos de repetição apresentam tamanho similar, porém seqüências distintas, assim como a seqüência à montante do ETS. Não foi determinado o número de blocos de repetição presente em cada cístron de L.(V.) braziliensis. Os estudos de ligação de fatores nucleares à região central do promotor mostraram que houve um reconhecimento diferencial dessa região entre a espécie homóloga L. (V.) braziliensis e as espécies heterólogas L. (V.) guyanensis, pertencente ao mesmo subgênero e L. (L.) amazonenis. Os complexos protéicos associados a essa região central apresentaram maior afinidade com a espécie heteróloga L. (V.) guyanensis. O fato de ser o ensaio de CAT trabalhoso e demorado levou à busca de outro repórter que evidenciasse a funcionalidade da região promotora ao mesmo tempo em que permitisse a quantificação dessa expressão, de modo a medir a força do promotor. Neste trabalho iniciamos a padronização para utilizar GFP como gene repórter no estudo da funcionalidade de promotores. Foi possível mostrar que a GFP é detectável em microscopia confocal e que as células que expressam GFP podem ser quantificadas em FACS, no entanto, essas detecções só foram possíveis em parasitas selecionados por uma marca de seleção, ou seja, numa transfecção estável. Assim, substituir o gene repórter CAT pelo gene GFP foi inadequado para os ensaios de transfecção transiente, impedindo que fosse medida a atividade da região promotora de L. (V.) braziliensis em diferentes espécies. / Leishmaniasis is the generic name of a serious public health problem that is caused by protozoa of the genus Leishmania. In its lifecycle, the parasite has two hosts, a vertebrate and an invertebrate. The genus Leishmania is divided into two subgenera: Leishmania (Leishmania) and Leishmania (Vianna), according to the position occupied by the promastigotes form at the gut of the invertebrate host. Genes that encode ribosomal RNA are exclusively transcribed by RNA polymerase I. The promoter region of the RNA polymerase I is known to present a species-specific recognition. Previous studies, based on transient expression assays with constructions in which the expression of the reporter gene CAT was directed by different regions of the promoter showed that RNA pol I promoter in Leishmania is species-specific, but in phylogenetically related species the expression of the reporter gene is higher than in the homologous species. The structural characterization of the promoter region of RNA pol I of L. (V.) braziliensis enabled to map the site of initiation of transcription and to define the promoter region of RNA pol I. The comparison of the determined 5\' region of the ETS region, with the corresponding region of other species of the subgenus L. (Leishmania), showed a high similarity both in size, as in the sequence of nucleotides. However, the IGS region and the repetitive blocks of nucleotides have similar size but different sequences. The studies of binding of nuclear factors to the central region of the promoter showed that there was a recognition between the species counterpart L. (V.) braziliensis and the heterologous species L. (V.) guyanensis, belonging to the same subgenus and L. (L.) amazonensis, that belongs to another subgenus. The complex protein associated with this central region showed greater affinity with the heterologous species L. (V.) guyanensis. The fact that to quantify the CAT expression represents a laborious and time consuming test lead us to search for another reporter that could identified the functionality of the promoter region and at the same time allowed the quantification of expression in order to measure the strength of the promoter. In the present work, we began the standardization for the use of GFP gene as reporter in the study of the functionality of promoters. It was possible to show that GFP is detectable in confocal microscope and the cells that express GFP can be quantified in FACS, however, these findings were made possible only in parasites selected by the mark, ie in a stable transfection. So, the replacement of CAT by GFP reporter showed to be inadequate for testing promoters in transient transfection. This prevented the measure of the activity of the promoter region of L. (V.) braziliensis in different species.
10

Expressão transcricional de ERFs em arroz submetidos ao estresse por ferro / Transcriptional Expression of ERFs in Rice Subjected to Iron Stress

Kruger, Mariana Madruga 19 December 2013 (has links)
Submitted by Gabriela Lopes (gmachadolopesufpel@gmail.com) on 2016-09-14T17:13:51Z No. of bitstreams: 2 license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) Mariana_Kruger.pdf: 976408 bytes, checksum: 6bfde814984b8524fba2bd3708136696 (MD5) / Approved for entry into archive by Aline Batista (alinehb.ufpel@gmail.com) on 2016-09-14T18:28:25Z (GMT) No. of bitstreams: 2 license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) Mariana_Kruger.pdf: 976408 bytes, checksum: 6bfde814984b8524fba2bd3708136696 (MD5) / Made available in DSpace on 2016-09-14T18:28:25Z (GMT). No. of bitstreams: 2 license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) Mariana_Kruger.pdf: 976408 bytes, checksum: 6bfde814984b8524fba2bd3708136696 (MD5) Previous issue date: 2013-12-19 / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior - CAPES / O arroz (Oryza sativa L.) é uma cultura que possui grande importância alimentar e econômica, sendo largamente cultivado em ecossistemas de terras baixas, o qual apresenta condições de drenagem deficiente. Nessas condições, o baixo potencial redox do meio leva a redução do ferro presente na solução do solo o qual torna-se prontamente disponível, podendo se tornar tóxico à planta quando absorvido em excesso. A toxidez por ferro é um dos mais importantes estresses abióticos a limitar a produção de arroz irrigado em nível mundial. Em áreas de cultivo de arroz do Rio Grande do Sul, o ferro tem causado problemas de toxidez e o uso de cultivares tolerantes torna-se uma importante estratégia para solucionar o problema, sendo necessário a identificação de genes frente aos diferentes mecanismos que conferem essa tolerância às plantas para aplicação em programas de melhoramento. Estudos recentes descobriram a importância de alguns fatores de transcrição responsivos ao etileno (ERFs) na resposta vegetal a diferentes estresses. Dessa forma o objetivo deste trabalho foi avaliar o perfil de expressão diferencial de 32 ERFs em arroz submetidos a condições de estresse por ferro, em três cultivares, Nipponbare, Epagri 108 (tolerante à toxidez por ferro) e BR-IRGA 409 (sensível à toxidez por ferro) e identificar elementos cis na região promotora de genes ERFs em arroz. Os resultados obtidos relatam que em arroz, a maioria dos membros da família ERF demonstram perfil diferencial nas cultivares Nipponbare, BR -IRGA 409 e Epagri 108, em condições de excesso de ferro. Os ERFs expressos na cultivar tolerante (Epagri 108) podem ser potencialmente importantes na indução da tolerância, enquanto que os que foram altamente induzidos na cultivar sensível (BR – IRGA 409) pertencem a um mecanismo de resposta o qual não é eficiente. Análises de elementos cis demonstram que a regulação da expressão de todos os ERFs analisados se dá de forma complexa e que existe uma associação entre a presença de determinado elemento cis com o perfil de expressão. / Rice (Oryza sativa L.) is a crop of high nourishment and financial importance, largely grown in flat and flooded ecosystems which present deficient drainage conditions. Under such conditions, the low redox potential of the environment leads to the reduction of iron present in the soil solution, which becomes readily available and can become toxic to the plant when absorbed in excess. Iron toxicity is one of the most important types of abiotic stress, limiting the production of irrigated rice worldwide. In rice growth areas of Rio Grande do Sul, Brazil, iron has caused toxicity problems and the use of tolerant cultivars becomes an important strategy in solving the problem, where the identification of genes is necessary in face of the different mechanisms which confer this tolerance to the plants for application in improvement programs. Recent studies discovered the importance of some transcription factors responsive to ethylene (ERFs) in vegetable response to different kinds of stress. Thus, the aim of this study was to evaluate the differential expression profile of 32 ERFs in rice subjected to iron stress conditions in three cultivars; Nipponbare, Epagri 108 (tolerant to iron toxicity) and Br-Irga 409 (sensitive to iron toxicity) and to identify cis elements in the ERF gene promotion region in rice. The results obtained demonstrate that in rice, most members of the ERF family showed differential profiles in the cultivars Npponbare, Epagri 108 and Br-Irga 409, under iron stress conditions. ERFs expressed in the tolerant cultivar (Epagri 108) may be potentially important in the induction of tolerance while those which were more highly induced in the sensitive cultivar (Br-IRGA 409) belong to a response mechanism which is not efficient. Analyses of cis elements demonstrate that the regulation of expression of all analyzed ERFs takes place in complex form and that there exists an association between the presence of a specific cis element and the expression profile.

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