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Item Avaliação da atividade da invertase de Saccharomyces cerevisiae imobilizada em polianilina sobre o caldo de cana(Universidade Federal de Goiás, 2010-02-19) BARBOSA, Eduardo Fernandes; FERNANDES, Kátia Flávia; http://lattes.cnpq.br/9737543228759171This work describes the immobilization of invertase on chemically synthesized polyaniline and activated with glutaraldehyde (PANIG) for production of invert syrup from sugarcane juice. Free invertase activity present in crude extract (E.B.) obtained from cells of Saccharomyces cerevisiae, was characterized for an evaluation of interferents present in the extract on enzyme activity (optimum conditions: temperature 50 ° C, pH of 4.5 in sodium acetate buffer 0.1 mol L-1 and reaction time of 10 minutes, with an activity of 11.31 ± 0.36 EU mL-1). We tested some parameters optimization of enzyme immobilization, such as amount of enzyme, immobilization time, pH and temperature of immobilization. The optimal immobilization was obtained in buffer sodium acetate 0.1 mol L-1 pH 4.5, immobilization time of 1 hour at 50°C and 169.55 EU mg-1 PANIG. The efficiency of immobilization was 0.86. The stability of the system PANIG-Invertase was tested against the storage time and thermostability, and after 75 days storage in buffer sodium acetate 0.1 mol L-1 pH 4.5 was obtained for 94% of initial activity with only 17% retained for the free enzyme. The immobilized invertase didn t change the optimal conditions compared to the free, but the immobilized was more stable in adverse conditions such as pH below and above optimum conditions showed an increase in thermostability. Some features of the hydrolysis product were evaluated (water activity, viscosity and color), compared to the sugarcane juice in nature, showing that the reactors allowed changes in sugarcane juice that expand the possibilities for using syrup obtained in the production of sweets, ice cream and syrups rich in fructose. The high stability of the system tested, along with its high retention of activity strongly suggests the use of the system in reactors.Item Análise de populações de Sclerotinia sclerotiorum em cultura de feijoeiro através de marcadores SSR.(Universidade Federal de Goiás, 2009-09-30) GOMES, Eriston Vieira; SILVA, Silvana Petrofeza da; http://lattes.cnpq.br/682399854496837379 Sclerotinia sclerotiorum isolates were collected under central pivot irrigations sistems, from Brazilian fields, divided in 4 populations (S2, S3, A and M) and analyzed to determine the genetic variability among and between populations using molecular markers based on microsatelittes. 10 primers were used and the amplification products were separate in poliacrilamida gels and the bands were silver stained. A total of 102 different haplotypes were identified, and the amount of haplotypes varied from 6 to 18 for locus. The genotypic diversity ranged from 65% to 91%. Analyses based on genetic diversity and fixation indices which indicate the variability between populations was 28.79% (FST = 28793) and the variability among populations was 71.21%. The Jaccard similarity index indicated that the populations S2 and A is genetically closer. The population S3 presented a similarity index of 0.44 compared with the populations S2 and A. The population M, originated from several collection sites, was considered genetically more distant showing a index of 0.46 compared with the others populations . The high variability between and among populations can indicate that, besides the possible introduction of new genotypes in the analyzed fields, could be having clonal and sexual reproduction in isolated of S. sclerotiorum from Brazilian cerrado.Item Caracterização Molecular Do Plasmídeo pLK39 Extraído De Uma Bactéria Endofítica Isolada De Solanum Lycocarpum(Universidade Federal de Goiás, 2002-02-28) OLIVEIRA, Vera Lúcia Cardoso de; BATAUS, Luiz Artur Mendes; http://lattes.cnpq.br/5637230378599476The characterization of the plasmid pLK39.was the aim of work.pLK39 was isolated from a endophytic Gram-negative, bactéria isolated from leaves form Solanum lycocarpum (Lobeira). In order to obtain a selection marker which would characterization in Escherichia coli the kanamycin, resistence gene from pUC4K. was subcloned into the PstIsite of pLK39. The characterization of PLK39 was basedon studies of stability, incompatibility, determination of the copy number and through DNA sequencing. The stability was carried out in Escherichia coli XL10 cells. Cells transformed with pLK39 were inoculated in Lúria broth containing Kanamycin (50μg/ml) during24 hours. After 24 hours of incubation one sample of the culture was in medim with and without selective pressure, and inoculated in Lúria broth without pressure médium, for 240 generations. After 240 generations 81,5% of cells maintained the plasmid, showing that pLK39is highly stable in Escherichia coli to make the incompatibility test, cells of Escherichia coli XL10 were co-transformed with pLK39/pUC18; pBR322 and pLK39/pACYC184. After 240 generations, the plasmid pLK39 iof detected in all systems used, showing the compatibility of pLK39 with the plasmids tested. The copý number pLK39 was estimated by the intensity of plasmidial bands in agarose gel, electrophoresis using a photodocumentation and analysis system (KODAK EDAS). The copý number of pLK39 was estimated as 25 copies per cell. PLK39 was digested with Sau3AI restriction and subcloned into the BamHI site of plasmid pUC18. The recombinant clones were sequenced and 1637 pb reading fragment was obtained. This sequence showed high homology with pSW200, pSW100, pEC3, pUCD5000 and pBERT, which were isolated from Gram-negative bacteria. This sequence possesses two possible genes. The ORF I showed high homology with mobB gene from plasmid pSW200 (96%), pSW100 (96%), pEC3 (94%), pUCD5000 (94%) and pBERT (87%). The ORF II showed homology with mobD gene from plasmids pSW200 (92%), pSW100 (90%), pEC3 (90%) and PUCD5000 (89%).Item Efeito da fonte de carbono e nitrogênio na produção de β 1,3 glucanases por Trichoderma asperellum(Universidade Federal de Goiás, 2008-05-29) SILVA, Regiane Christine da; ULHOA, Cirano Jose; http://lattes.cnpq.br/8368469162867277The β-1.3-glucanases have several functions in the cell as fungal metabolic in hydrolysis of polysaccharides for possible cellular assimilation; morphogenetic function in the hydrolysis or modifications of the cell wall, which includes growth and extension of the wall, altering the structure and composition of the wall and autolysis. In addition to presenting ecological importance, since these enzymes are involved in biological control, through the process of mycoparasitism. The gender Trichoderma comprises a group of filamentous fungi, saprophytic soil, found on decomposing organic matter in the rhizosphere, and some plants. Considering the importance of β-1.3 glucanases proposing in this study to evaluate the production of β-1.3 - glucanases by T. asperellum using different sources of carbon and nitrogen. The enzyme was produced using cell wall of Rizoctonia solani (PCRS), chitin, chitosan, starch, cellulose, sucrose, maltose, lactose, celobiose and glucose. The best production of enzymes was obtained in the middle containing PCRS, with two bands of activity of polyacrylamide gel were found. Furthermore, the effect of varying concentrations of ammonium sulfate (2, 4, 6, 8, 20, 40 and 60 mM) in the production of β-1.3-glucanase was evaluated during the growth of the fungus in PCRS. A great increase in activity of the enzyme was detected in the presence of nitrogen source. We can see that there was a considerable increase in the expression of the enzyme of low molecular weight at this culture conditions.Item Construção e análise de uma biblioteca de cDNA de Trichoderma harzianum crescido na presença de parede celular de Fusarium solani(Universidade Federal de Goiás, 2010-02-26) STEINDORFF, Andrei Stecca; SILVA, Roberto do Nascimento; http://lattes.cnpq.br/5771309284349109; ULHOA, Cirano Jose; http://lattes.cnpq.br/8368469162867277Species of Trichoderma are commercially applied as biological control agents against plant fungal pathogens. Their action is based on a set of events, such as the production of antifungal metabolites, competition for space and nutrients and mycoparasitism. However the biocontrol mechanism of the interaction between Trichoderma harzianum and Fusarium solani is not yet understanded at the transcriptome level. This study aimed to initiate the preliminary development of an expressed sequence tag (EST) database for Trichoderma harzianum and thereby gain potentially useful information on Trichoderma gene sequences in order to elucidate the integrated biocontrol mechanism. Partial sequencing of anonymous cDNA clones is a widely used technique for gene identification. A directional cDNA library has been constructed from mycelium of Trichoderma harzianum grown on cell wall isolated from Fusarium solani. 3984 clones have been randomly selected, subjected to single-pass sequencing from the 5 end of the vector, and identified by sequence similarity searches against gene sequences in GenBank fungal database. Of the 3984 mycelium clones, 40.8% exhibit similarity to known genes. Analysis of the identified clones indicated sequence similarity to a broad diversity of genes encoding proteins such as enzymes, structural proteins, and regulatory factors. A significant proportion of genes identified were involved in processes related to mycoparasitism (3,81%), as would be expected in biocontrol fungus. These results present the successful application of EST analysis in the interaction of Trichoderma harzianum and Fusarium solani to provide a preliminary indication of gene expression.