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dc.creatorSoares, Isis Capella-
dc.creator.Latteshttp://lattes.cnpq.br/4206441276341228por
dc.contributor.advisor1Araújo, Jean Luiz Simões de-
dc.contributor.advisor1Latteshttp://lattes.cnpq.br/4344808401973010por
dc.contributor.advisor-co1Pacheco, Rafael Sanches-
dc.contributor.advisor-co1Latteshttp://lattes.cnpq.br/2878448197446930por
dc.contributor.referee1Pacheco, Rafael Sanches-
dc.contributor.referee1Latteshttp://lattes.cnpq.br/2878448197446930por
dc.date.accessioned2022-06-01T13:08:51Z-
dc.date.issued2019-12-18-
dc.identifier.citationSOARES, Isis Capella. Avaliação da população de duas espécies diazotróficas associativas em tecidos de braquiária e milho utilizando PCR quantitativa. 2019. 85 f. Dissertação (Mestrado em Fitossanidade e Biotecnologia Aplicada). Instituto de Ciências Biológicas e da Saúde, Universidade Federal Rural do Rio de Janeiro, Seropédica, 2019.por
dc.identifier.urihttps://tede.ufrrj.br/jspui/handle/jspui/5721-
dc.description.resumoBactérias diazotróficas dos gêneros Azospirillum e Herbaspirillum, em associação com gramíneas como o milho e forrageiras do gênero Brachiaria podem promover efeitos benéficos às plantas, como a fixação biológica de nitrogênio (FBN). A técnica de PCR quantitativa (qPCR) é eficiente no monitoramento de populações de bactérias diazotróficas inoculadas, porém sua aplicabilidade em nível de estirpe necessita de maiores estudos. Este trabalho teve o objetivo de selecionar oligonucleotídeos (primers) para posterior quantificação da população de A. brasilense em tecidos de raiz e colmo de braquiária (Brachiaria spp.) e H. seropedicae em raiz de milho (Zea mays) por qPCR. Os primers desenhados foram avaliados quanto a sua especificidade em nível de estirpe e espécie, por meio de PCR convencional. Já a sensibilidade e eficiência dos primers foram determinadas por reações de qPCR. A quantificação por qPCR estirpeespecífica de A. brasilense e H. seropedicae associadas a tecidos de braquiária e milho foi feita a partir de duas curvas-padrão diferentes. Os resultados obtidos por PCR quantitativa com os pares de primer selecionados foram comparados a contagem por microgota. A quantificação da estirpe A. brasilense Sp245 foi feita a partir do DNA extraído de tecidos de colmo e raiz de cultivares de braquiária inoculada e sem inoculação, crescidas em condições de campo. A população bacteriana das estirpes A. brasilense Sp245 e H. Seropedicae ZAE94 foi quantificada através do DNA genômico de raiz de milho inoculado com cada estirpe-alvo e plantas controle, sem inoculação, crescidas em condições de casa de vegetação com substrato estéril sob duas dosagens de N (3 e 0,3 mM). O par de primer Sp245p10, foi específico para a estirpe Sp245 e os pares de primer ZAEF1R1 e ZAEF2R2 foram específicos para a estirpe ZAE94. A quantificação, por qPCR, das estirpes Sp245 e ZAE94 apresentou resultados similares à contagem por microgota. Em relação ao experimento a campo com braquiária, o número de células bacterianas da estirpe Sp245 foi maior em plantas inoculadas das cultivares Basilik e Piatã. No experimento com milho em casa de vegetação, a dose de 3 mM de N favoreceu a população endógena de bactérias da estirpe Sp245 em plantas do tratamento controle, enquanto que a dose de N não interferiu na população das estirpes Sp245 e ZAE94 nas plantas inoculadaspor
dc.description.abstractDiazotrophic bacteria, from Azospirillum and Herbaspirillum genus, in association with grasses, such as maize and brachiaria, can promote benefits for the plant, such as biological nitrogen fixation (BNF). Quantitative PCR technique (qPCR) is effective in monitoring the endophytic bacterial population, however its applicability at strainspecific level needs further study. This study aimed to select oligonucleotides (primers) for quantification of strains populations of A. brasilense in roots and stems of brachiaria (Brachiaria spp.) and H. seropedicae in root of maize (Zea mays) by strain-specific qPCR. The primers designed were evaluated by conventional PCR for specificity for strain and species. Primers sensitivity and efficiency were determined by qPCR reactions. The quantification by strain-specific qPCR of the A. brasilense and H. seropedicae associated with brachiaria and maize tissues were made by using two different standard curves. The results of quantitative PCR using the selected primers were compared to the microdrop count technique. The strain A. brasilense Sp245 was quantified from DNA extracted from stems and root tissues of inoculated and noninoculated brachiaria cultivars grown under field conditions. The bacterial population of the A. brasilense Sp245 and H. seropedicae ZAE94 strains was quantified by maize root genomic DNA from inoculated and non-inoculated plants. Maize plants were grown under greenhouse conditions with sterile substrate and different N doses (3 and 0.3 mM). The primer Sp245p10 was specific for Sp245 strain, and the primers ZAEF1R1 and ZAEF2R2 were specific for the ZAE94 strain. Quantifications by qPCR of Sp245 and ZAE94 strains presented similar results to micro drop count. In the field-grow brachiaria experiment, the number of Sp245 strain cells was higher in inoculated plants from Basilik and Piatã cultivars. In the greenhouse experiment, the N dose with 3 mM favored the endogenous population of Sp245 bacteria in non-inoculated maize plants, while the N dose did not affect the Sp245 and ZAE94 population in inoculated plantseng
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dc.description.sponsorshipCAPES - Coordenação de Aperfeiçoamento de Pessoal de Nível Superiorpor
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dc.languageporpor
dc.publisherUniversidade Federal Rural do Rio de Janeiropor
dc.publisher.departmentInstituto de Ciências Biológicas e da Saúdepor
dc.publisher.countryBrasilpor
dc.publisher.initialsUFRRJpor
dc.publisher.programPrograma de Pós-Graduação em Fitossanidade e Biotecnologia Aplicadapor
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Azospirillum brasilense promotes increases in growth and nitrogen use efficiency of maize genotypes. PLoS ONE, v. 14, n. 4, p. 1–19, 2019. ZHANG, T.; FANG, H. H. P. Applications of real-time polymerase chain reaction for quantification of microorganisms in environmental samples. Applied Microbiology and Biotechnology, v. 70, n. 3, p. 281–289, 2006.por
dc.rightsAcesso Abertopor
dc.subjectAzospirillum brasilensepor
dc.subjectHerbaspirillum seropedicaepor
dc.subjectstrain-specific primerpor
dc.subjectReal time PCRpor
dc.subjectAzospirillum brasilenseeng
dc.subjectHerbaspirillum seropedicaeeng
dc.subjectstrain-specific primereng
dc.subjectReal time PCReng
dc.subject.cnpqBiologia Geralpor
dc.titleAvaliação da população de duas espécies diazotróficas associativas em tecidos de braquiária e milho utilizando PCR quantitativapor
dc.title.alternativePopulation evaluation of two associated diazotrophic species in brachiaria and maize tissues using quantitative PCReng
dc.typeDissertaçãopor
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