Programa de Pós-graduação em Ciências Biológicas
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Navegando Programa de Pós-graduação em Ciências Biológicas por Por Orientador "COELHO, Alexandre Siqueira Guedes"
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Item Avaliação de critérios de compatibilidade entre pares de primers para otimização de sistemas multiplex de genotipagem(Universidade Federal de Goiás, 2010-01-12) BARBOSA, Ana Clara de Oliveira Ferraz; COELHO, Alexandre Siqueira Guedes; http://lattes.cnpq.br/0840926305216925The progress of Molecular Biology and Genetics provided the appearance of several molecular markers that detect the genetic polymorphism directly at DNA. Among these markers are the microsatellites (SSR), which are distinguished by their high degree of polymorphism. The use of these markers for individual genotyping has evolved into multiplex systems, which allow many SSR fragments to be detected and analyzed simultaneously. Currently there are several articles in literature discussing the criteria to be used in the primer design for use in PCR, as well as various softwares are available for this end. However, there are few studies and tools for the analysis of compatibility between pairs of primers for use in multiplex systems, where multiple fragments are simultaneously amplified using PCR. This paper evaluated different criteria for compatibility between pairs of primers. A set of 74 combinations of pairs of primers, involving the amplification of 94 SSR loci were evaluated in duplex systems. The same combinations were evaluated according to different criteria, including the degree of complementarity between primers, the magnitude of differences of denaturation temperatures (Tm) and the tendency to annealing between pairs of primers based on the Gibbs free energy resulting from the association between them. The comparison between the different criteria allowed the identification of a set of criteria with positive predictive value equal to 94%. These criteria were implemented for use in a software called Multiplexer, which from the analysis in sequence of pairs of primers, suggests compatible combinations for use in multiplex genotyping systems. Using this tool can significantly reduce the costs related to laboratory activities for genotyping using PCR.Item Anotação e caracterização preliminar de genes de celulose sintase em diferentes espécies de Eucalyptus(Universidade Federal de Goiás, 2006-12-01) SALAZAR, Marcela Mendes; COELHO, Alexandre Siqueira Guedes; http://lattes.cnpq.br/0840926305216925Cellulose is the world s most abundant polymer, being the main constituent of plant biomass. Genes from CesA family, that encodes the cellulose synthase enzyme, was identified in several plant species, especially in Arabidopsis thaliana, in which three of them (AtCesA4, AtCesA7 e AtCesA8) were associated with secondary cell wall cellulose production. Eucalyptus species represent an important target of genetic improvement studies, since it is the most planted forest genus in the world, beyond deserving a special place in the international market of cellulose and paper. The molecular breeding technology enables that gene characterization can be applied to forest species genetic improvement programs with the aim to improve the quality and productivity of its products. In this work, 320 Eucalyptus ESTs, separated in four genes related with cellulose production in secondary cell wall, using AtCesA4, AtCesA7 e AtCesA8 genes as reference. For these genes, primers were designed in order to screen an Eucalyptus BACs library. An emphasis was given to genes that are orthologs to AtCesA7 from Arabidopsis thaliana for witch it was sequenced a 1197 base pairs region from the BACs library and this served as a support to expression level studies of this genes in different species/tissues, showing that this is preferentially expressed in xylem and weakly expressed in leaves. The expression level of this gene was higher in E. urophylla than in other species studied. Sequences from approximately 500bp was obtained from different Eucalyptus species and in these, with one intron between two exons, the amount of SNPs in the intron (4), as waited, was higher than that found in exons (1 in each), although the intron nucleotide diversity index (π) (0,0824) were less than in the exon (0,2029). In this manner, one expects that this work can contribute for one better understanding of the mechanisms involved in biosynthesis and regulation of the cellulose pathway in Eucalyptus species, as well as subsidize genetic mapping studies and linkage disequilibrium analysis for this genus.