89 resultados para suppression subtractive hybridization (SSH)

em Repositório Institucional UNESP - Universidade Estadual Paulista "Julio de Mesquita Filho"


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Late leaf spot (LLS), caused by the fun.-us Cercosporidium personatum, is one of the most severe diseases in peanut (Arachis hypogaea). The vast majority of commercial cultivars do not exhibit satisfactory levels of resistance to the pathogen, whereas non-commercial genotypes cv. 850 and cv. 909 are resistant to LLS and show symptoms similar to hypersensitive response (HR) lesions. In the present study, we investigated the molecular components of the initial stages of the resistance by gene expression profiling using suppression subtractive hybridization and differential screening of cDNA macroarray techniques. Gene expression analyses have allowed us to identify more than 700 peanut unique expressed sequence taus (EST) involved in several aspects of the early stages of C. personatum pathogenesis, such as components of defense signaling pathways, gene expression regulators, cell cycle controlling genes and components of the biosynthesis of transducer and antimicrobial compounds. The most significantly induced gene corresponds to a novel O'-methyltranferase, suggesting its involvement in the production of local lesions in C. personatum-resistant A. hypogea genotypes. Taken together, our results contribute to elucidate the defense strategies of peanut and provide the framework for the generation of pathogen-resistant peanut cultivars. (C) 2007 Elsevier B.V. All rights reserved.

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Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

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Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

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Pós-graduação em Genética - IBILCE

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Cervical cancer is the second most frequent cancer in women worldwide and is associated with genetic alterations, infection with human papilloma virus (HPV), angiogenesis and inflammatory processes. The idea that inflammation is involved in tumorigenesis is supported by the frequent appearance of cancer in areas of chronic inflammation. On the other hand, the inflammatory response is controlled by the action of anti-inflammatory mediators, among these mediators, annexin A1 (ANXA1), a 37 kDa protein was detected as a modulator of inflammatory processes and is expressed by tumor cells. The study was carried out on the epithelial cancer cell line (SiHa) treated with the peptide of annexin A1 (ANXA1Ac2-26). We combined subtraction hybridization approach, Ingenuity Systems software and quantitative PCR, in order to evaluate gene expression influenced by ANXA1. We observed that ANXA1Ac2-26 inhibited proliferation in SiHa cells after 72h. In these cells, 55 genes exhibited changes in expression levels in response to peptide treatment. Six genes were selected and the expression results of 5 up-regulated genes (TPT1, LDHA, NCOA3, HIF1A, RAB13) and one down-regulated gene (ID1) were research by real time quantitative PCR. Four more genes (BMP4, BMPR1B, SMAD1 and SMAD4) of the ID1 pathway were investigated and only one (BMPR1B) shows the same down regulation. The data indicate the involvement of ANXA1Ac2-26 in the altered expression of genes involved in tumorigenic processes, which could potentially be applied as a therapeutic indicator of cervical cancer.

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Objetivou-se estudar a influência de diversas concentrações do meio MT, sacarose, vitaminas, carvão ativado e ácido giberélico no cultivo de embriões imaturos oriundos do cruzamento entre laranjeira 'Pêra Rio' x tangerineira 'PONCÃ' . Os embriões foram excisados sob condições assépticas e inoculados em 15 mL do meio de cultura MT, de acordo com cada experimento a seguir: 1) concentrações do meio de cultura MT (0%, 50%, 100%, 150% e 200%) combinados com 0, 30, 60 e 90 g.L-1 de sacarose; 2) concentrações de vitaminas do meio MT (0%, 50%, 100%, 150% e 200%) combinados com 0, 30, 60 e 90 g.L-1 de sacarose e; 3) concentrações de carvão ativado (0; 0.5; 1; 1.5 e 2 g.L-1) combinados com GA3 (0; 0.01; 0.1; 1 e 10 mg.L-1). Após a inoculação, os embriões foram mantidos por 90 dias em sala de crescimento à temperatura de 27±1ºC, fotoperíodo de 16 horas e irradiância de 32 µmol.m-2.s-1. A utilização de 50% e 100% do meio MT associado a 60 e 90 g.L-1 de sacarose, respectivamente, acrescido de 0.01 mg.L-1 de GA3, proporcionou melhor desenvolvimento de embriões globulares. Não houve necessidade da adição de carvão ativado e vitaminas no meio MT para o cultivo de embriões globulares.

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The electrical characterization of a high efficient multilayer polymer light emitting diode using poly[(2-methoxy-5-hexyloxy)-p-phenylenevinylene] as the emissive layer and an anionic fluorinated surfactant as the electron transport layer was performed. For the sake of comparison, a conventional single layer device was fabricated. The density current vs. voltage measurements revealed that the conventional device has a higher threshold voltage and lower current compared to the surfactant modified device. The effective barrier height for electron injection was suppressed. The influence of the interfaces and bulk contributions to the dc and high frequencies conductivities of the devices was also discussed. (c) 2006 Springer Science + Business Media, Inc.

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The local and systemic production of prostaglandin E-2 (PGE(2)) and its actions in phagocytes lead to immunosuppressive conditions. PGE2 is produced at high levels during inflammation, and its suppressive effects are caused by the ligation of the E prostanoid receptors EP2 and EP4, which results in the production of cyclic AMP. However, PGE(2) also exhibits immunostimulatory properties due to binding to EP3, which results in decreased cAMP levels. The various guanine nucleotide-binding proteins (G proteins) that are coupled to the different EP receptors account for the pleiotropic roles of PGE(2) in different disease states. Here, we discuss the production of PGE(2) and the actions of this prostanoid in phagocytes from different tissues, the relative contribution of PGE(2) to the modulation of innate immune responses, and the novel therapeutic opportunities that can be used to control inflammatory responses.

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The piezoelectric quartz crystal resonators modified with oligonucleotide probes were used for detection of hepatitis C virus (HCV) in serum. The gold electrodes on either rough or smooth surface crystals were modified with a self-assembled monolayer of cystamine. After activation with glutaraldehyde, either avidin or streptavidin were immobilized and used for attachment of biotinylated DNA probes (four different sequences). Piezoelectric biosensors were used in a flow-through setup for direct monitoring of DNA resulting from the reverse transcriptase-linked polymerase chain reaction (RT-PCR) amplification of the original viral RNA. The samples of patients with hepatitis C were analyzed and the results were compared with the standard RT-PCR procedure (Amplicor test kit of Roche, microwell format with spectrophotometric evaluation). The piezoelectric hybridization assay was completed in 10 min and the same sensing surface was suitable for repeated use. (C) 2004 Elsevier B.V. All rights reserved.

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A new electrochemical hybridization biosensor protocol without an external indicator is described. The biosensor format involves the immobilization of inosine-substituted (guanine-free) probe onto the carbon paste transducer, and a direct chronopotentiometric detection of the duplex formation by the appearance of the guanine oxidation peak of the target. Such a use of the intrinsic DNA electrochemical response for monitoring hybridization events offers several advantages (over the common use of external indicators), including the appearance of a new peak, a Aat background, or simplicity. A 4 min short hybridization period allows a detection limit around 120 ng/ml. Performance characteristics of the sensor are described along with future prospects. (C) 1998 Elsevier B.V. B.V. All rights reserved.