Microbiologia

URI permanente desta comunidadehttps://locus.ufv.br/handle/123456789/11840

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2002 - 20054

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Autor: search.filters.author.Pereira, Jorge FernandoData final: 2007

Resultados da Pesquisa

Agora exibindo 1 - 4 de 4
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    Development of a transformation system for Penicillium brevicompactum based on the Fusarium oxysporum nitrate reductase gene
    (Brazilian Journal of Microbiology, 2005-04) Queiroz, Marisa Vieira de; Pereira, Jorge Fernando; Ribeiro, Ronney Adriano; Soares, Marcos Antônio; Ribeiro, João Batista; Araújo, Elza Fernandes de; Varavallo, Maurílio Antônio
    Penicillium brevicompactum is a filamentous fungus that presents a potential for industrial use due its efficient pectinase production. A heterologous transformation system was developed for P. brevicompactum based on the complementation of a nitrate reductase mutant. Nitrate reductase mutants were obtained by resistance to chlorate in a rate of 23.24% when compared to other mutations that lead to the chlorate resistance. One mutant named 4457-18X was chosen for the transformation experiments with the pNH24 vector containing de Fusarium oxysporum nitrate reductase gene. A frequency of approximately 3 transformants/µg DNA was obtained using the circular vector pNH24. This frequency was multiplied about 10 fold using the linearized vector with the Xba I restriction enzyme. Southern analysis of the transformants showed a tendency of the linearized vector to diminish the number of integrations compared to the use of the circular vector. The integration was random and stable in the analyzed transformants. The establishment of a transformation system for P. brevicompactum is fundamental for genetic manipulation of this microorganism.
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    Characterization, regulation, and phylogenetic analyses of the Penicillium griseoroseum nitrate reductase gene and its use as selection marker for homologous transformation
    (Canadian Journal Of Microbiology, 2004) Pereira, Jorge Fernando; Queiroz, Marisa Vieira de; Lopes, Francis Júlio Fagundes; Rocha, Rodrigo Barros; Daboussi, Marie-Josée; Araújo, Elza Fernandes de
    Penicillium griseoroseum has been studied because of its efficient pectinases production. In this work, the Penicillium griseoroseum nitrate reductase gene was characterized, transcriptionally analyzed in different nitrogen sources, and used to create a phylogenetic tree and to develop a homologous transformation system. The regulatory region contained consensus signals involved in nitrogen metabolism and the structural region was possibly interrupted by 6 introns coding for a deduced protein with 864 amino acids. RT-PCR analysis revealed high amounts of niaD transcript in the presence of nitrate. Transcription was repressed by ammonium, urea, and glutamine showing an efficient turnover of the niaD mRNA. Phylogenetics analysis showed distinct groups clearly separated in accordance with the classical taxonomy. A mutant with a 122-bp deletion was used in homologous transformation experiments and showed a transformation frequency of 14 transformants/microg DNA. All analyzed transformants showed that both single- and double-crossover recombination occurred at the niaD locus. The establishment of this homologous transformation system is an essential step for the improvement of pectinase production in Penicillium griseoroseum.
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    Molecular characterization and evaluation of pectinase and cellulase production of Penicillium spp.
    (Biotechnology Letters, 2002-05) Pereira, Jorge Fernando; Queiroz, Marisa Vieira de; Gomes, Eliane Aparecida; Muro-Abad, Júpiter Israel; Araújo, Elza Fernandes de
    Penicillium species were analyzed with molecular markers and for pectinase and cellulase production. RAPD and PCR-RFLP analysis indicated high polymorphism among at least 5 of 10 Penicillium species. Five species were chosen for pectinase and cellulase production in liquid medium and four of which appeared similar based on molecular analyses. P. brevicompactum and P. griseoroseum gave the highest pectinase production and were highly divergent by molecular techniques.
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    Development of a transformation system for Crinipellis perniciosa, the causal agent of witches' broom in cocoa plants
    (Current Genetics, 2002-12-17) Lima, Juliana Oliveira; Santos, Jildete Karla dos; Pereira, Jorge Fernando; Resende, Mário Lúcio Vilela de; Araújo, Elza Fernandes de; Queiroz, Marisa Vieira de
    Protoplasts of the pathogenic plant fungus, Crinipellis perniciosa, were transformed to hygromycin B resistance using the pAN7-1 plasmid, which contains the Escherichia coli hph gene under the control of Aspergillus nidulans regulatory sequences. The pAN7-1 plasmid was introduced by PEG/CaCl2 treatment. Transformation frequencies of 1.6–2.5 transformants/μg of DNA were achieved. About 54% of the transformants were abortive and 40 analyzed transformants were mitotically stable and showed different hygromycin B resistance levels. The presence of the hph gene was checked by PCR in five transformants and the integration of multiple plasmid copies into different genome sites was observed by Southern analysis. This is the first report of a C. perniciosa transformation system and represents an important step for further research into genetic manipulation of this fungal plant pathogen.