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Item Métodos diagnósticos para o vírus mayaro: revisão sistemática e avaliação molecular em pacientes arbovirose like em unidade municipal de sáude de Goiânia-Goiás(Universidade Federal de Goiás, 2020-02-17) Abrantes, Gabrielly Regis; Anunciaçāo, Carlos Eduardo; http://lattes.cnpq.br/4354412874919580; Lacerda, Elisângela de Paula Silveira; http://lattes.cnpq.br/9390789693192751; Lacerda, Elisângela de Paula Silveira; Anunciaçāo, Carlos Eduardo; Lino, Guilherme; Mrué, FátimaDiseases caused by viruses are considered an often neglected public health problem that affects thousands of people worldwide, every year. Symptoms like fever, arthralgia and rash. classify patients as syndromic dengue or arbovirus like. The differential diagnosis of arbovoresis is by serological and molecular techniques. The present study aimed to evaluate diagnostic techniques in studies published in the literature in order to develop a Systematic Literature Review on the detection of the Mayaro virus that allows health professionals to carry out an efficient laboratory diagnosis, as well as the application of molecular tests for the diagnosis of MAYV. The available literature was evaluated by systematic review, focusing on the diagnosis of the Mayaro virus in humans and its methodologies. The systematic review was carried out through an exploratory study in the databases: Scielo (Scientific Eletronic Library Online), Medline (Medical Literature Analysis and Retrieval System Online), Scopus database, PubMed, LILACS from June 8, 2019 to 02 November 2019, selecting scientific articles according to the criteria of the PRISMA (2009) recommendation. The data showed that the circulation of the Mayaro virus in urban regions is already well established worldwide and that the misdiagnosis is frequent due to similar symptoms with other arboviruses. There is a wide use of varied methods over time in the diagnosis for this arbovirus, requiring further research to implement a standardized, sensitive and specific method. In a second stage of the research, between May to August 2017 and January to June 2018, 452 samples of patients with syndromic dengue symptoms (arboviruses like) were screened. The samples were sent to the Molecular Genetics and Cytogenetics Laboratory of the Federal University of Goiás, where they were subjected to molecular screening by Polymerase Chain Reaction followed by Reverse Transcriptase (RT-PCR) reaction. All positive cases for the first PCR were confirmed on RT-qPCR for viral detection. Thirteen positive samples went through both tests and thus were subjected to Sanger sequencing. In this experimental study, 17 samples were positive for the Mayaro virus infection (RT-PCR and RT-qPCR), and co-infections with the Dengue virus were also found by molecular screening carried out in parallel for the Dengue virus by RT-PCR and RT-qPCR. The positive samples for MAYV / DENV were analyzed phylogenetically, suggesting circulation of Dengue serotype 2 and a Peruvian strain of Mayaro Virus (strain IQ 4235) in Goiânia.Item Estudo filogeográfico de Micrurus lemniscatus (LINNAEUS, 1758) (SERPENTES: ELAPIDAE)(Universidade Federal de Goiás, 2014-03-10) Abreu, Tatianne Piza Ferrari; Silva Junior, Nelson Jorge da; http://lattes.cnpq.br/6544526824923185; Collevatti, Rosane Garcia; http://lattes.cnpq.br/9979596352166630; Collevatti, Rosane Garcia; Maciel, Natan Medeiros; Vaz-Silva, WilianMicrurus lemniscatus is a South American coral snake species, popularly known as “coral verdadeira”. It is widely distributed in Seasonally Dry Forests (SDF), Gallery Forests and Rainforests. The Tertiary events and the climatic oscillations of the Quaternary affected the distribution of these ecosystems altering, in turn, the distribution of animals associated to these habitats. We hypothesize that the forest expansion and contraction cycles caused by climate fluctuations may have influenced the current distribution and genetic structure of M. lemniscatus. This study aimed to study the evolutionary relationships and patterns of divergence among M. lemniscatus lineages and infer the historical biogeographic events that influenced the distribution and genetic variation. Twenty-nine individuals of M. lemniscatus were sampled from 16 localities in the states of Tocantins, Bahia, Goiás, Alagoas, Mato Grosso, Maranhão, Pará and Amazônia. Three mitochondrial regions (COI, 16S and ND4L) were sequenced, and together generated a fragment of 1595 bp and 23 different haplotypes. The analyses showed a very ancient lineage divergence ~ 4.5 Myr and high genetic differentiation among localities (FST=0.932; p<0.01), suggesting a limited gene flow among geographical regions. The demographic analyzes and neutrality tests indicated that there is no sign of expansion and that populations have constant size (Tajima’s D = 0.521; p = 0.763 and Mismatch distribution = 0.009; p= 0.779). In the analysis of the evolutionary relationship between haplotypes (by median-joining network method), no relationship between lineage and geographical space was found, suggesting incomplete lineage sorting. The results corroborate and give evidence that populations of M. lemniscatus were distributed in a more continuous region in the past, and the current distribution of this species may be the result of the reduction and separation of the geographical scope of their distribution, rather than having itself been an expansion event. This may be the result of cycles expansion of SDF and retraction of the rainforests during the cool and dry phases of the Quaternary.Item Bioprospecção de bactérias quitinolíticas e caracterização da atividade da enzima quitinase(Universidade Federal de Goiás, 2018-04-12) Alexandre, Artur Ribeiro de Sá; Amaral, André Corrêa; http://lattes.cnpq.br/8801299423520104; Amaral, André Corrêa; Carneiro, Lilian Carla; Vieira, José Daniel GonçalvesBioprospecting is defined as the search for organisms, genes, enzymes, compounds, processes and any pieces from living beings, that could have economic potential, and eventually lead to a product development. Thus, enzymes, protein catalysts, have aroused the interest of industries for its conversion mode specific biomass, low energy cost and not production of toxic waste. Chitinases are enzymes that catalyze the lysis of chitin (biopolymer composed of N-acetylglucosamine monomers), and thus have several applications, among them: obtaining N-acetylglucosamine monomers, used in the production of prebiotics; degradation of chitinous waste from fishing activities; and the use in control of fungi and insects. The present work aimed to bioprospect chitinase producing bacteria from soil samples and to characterize the enzymatic activity patterns of the best bacterial isolate. To do so, a screening with 17 soil samples collected in the states of Minas Gerais, Santa Catarina and Rio Grande do Sul, was carried out using a culture medium with colloidal chitin. Thirteen chitinase producing bacteria were obtained, among them, the isolate Q1 (identified as Paenibacillus illinoisensis), demonstrated to be a good producer of the enzyme and thus was selected for determination and optimization of its chitinase activity evaluating reaction time, temperature and pH. The chitinase produced by the isolate showed 0.098 U of activity, which was subsequently improved to 0.66 U when tested under the optimal conditions of 1 hour of reaction at 37 ºC and pH 4, an increase of 573% over the initial value. The values of chitinase activity from the isolate P. illinoisensis are close to those found in other studies, which also emphasize the potential application of the enzyme, mainly in the control of phytopathogenic pests. Bioprospecting of chitinase producing bacteria is possible and promising.Item Análise de polimorfismos em genes de detoxificação e reparo em indivíduos de municípios goianos(Universidade Federal de Goiás, 2017-03-10) Alves, Alessandro Arruda; Silva, Daniela de Melo e; http://lattes.cnpq.br/9895211901348365; Soares, Thannya Nascimento; Rodrigues, Flavia MeloIn Brazilian agriculture, the State of Goiás is one of the largest producers of corn, beans, cotton, sorghum, sugarcane and soybeans, with 75% of its exports composed of products related to agriculture and minerals. Thus, a large part of the economy of the State of Goiás is focused on rural activities, justifying the intense use of pesticides in the region. These pesticides will be metabolized in the liver in two main phases, the first one of bioactivation converting the original substances into more polar metabolites, through oxidation reduction reactions, hydrogenation and dehalogenation, most of the reactions are made by the cytochrome P450 (CYP) superfamily And the second phase in which conjugation reactions occur, in which metabolites are converted into non-toxic products. Agrochemicals are also known to cause various human health damage, such as cancer, cholinergic syndrome, Parkinson's disease and Alzheimer's as well as cytogenetic changes such as DNA damage and chromosome translocations. Thus, polymorphisms in enzymes acting in the path of detoxification and DNA repair may be important biomarkers of diseases in individuals exposed to pesticides. The present study sequenced 3 regions of the following genes: CYP2E1 (promoter region), CYP1A1 (exon 7) and OGG1 (intron region) of 260 individuals from Goiânia and Aparecida de Goiânia, GO. 20 ml of blood was collected, DNA was extracted using commercial kits and PCR was performed. Subsequently, the amplicons were sequenced by the automated Sanger method and compared to the 1000 Genome project. The population of this study was composed of 159 men and 101 women, mean age of 38 years. A total of 5 SNP's were found in the Goias population, being rs2031922 (change of C> T and frequency of the T allele of 0.05) of the CYP2E1 gene, rs1048943 (change of A> G and frequency of the G allele of 0.1 ) And rs1799814 (change of C> A and frequency of allele A of 0.07) of the CYP1A1 gene and rs1052133 (change of C> G and frequency of allele G of 0.2) and rs293795 (change of A> G and frequency Of the 0.18 T allele) of the OGG1 gene. All loci did not deviate from the Hardy-Weinberg equilibrium, nor was a statistically significant difference between age and genotype composition. The rs1048943 and rs1799814 loci of the CYP1A1 gene showed a moderate disequilibrium linkage and the rs1052133 and rs293795 loci of the OGG1 gene showed a strong disequilibrium linkage. The haplotype inference revealed a frequency of the haplotypes of loci rs1048943 and rs1799814 of A / C = 0.8, G / A = 0.12, A / A = 0.07 of the CYP1A1 gene. The haplotypes of the loci rs293795 and rs1052133 had A / G = 0.07, A / C = 0.8, G / C = 0.13. Due to the important participation in detoxification or DNA repair and relationship with pesticides, SNPs rs2031920 of the CYP2E1 gene, rs1048943 of CYP1A1 and rs1052133 of the OGG1 gene are potential biomarkers of susceptibility to disease development in individuals who have contact with these xenobioticsItem Avaliação molecular dos polimorfismos genéticos na susceptibilidade à nefropatia diabética por alterações hemodinâmicas e endoteliais(Universidade Federal de Goiás, 2018-12-19) Anjos, Laura Raniere Borges dos; Santos, Rodrigo da Silva; http://lattes.cnpq.br/4806187026900959; Reis, Angela Adamski da Silva; http://lattes.cnpq.br/3243656364470085; Cruz, Aparecido Divino da; Cruz, Aline Helena da Silva; Rocha, Thiago Lopes; Ternes, Yves Mauro FernandesDiabetic nephropathy (DN) is the major microvascular complication of Diabetes mellitus (DM) and is among the leading causes of kidney disease. This pathology is important because of glomerular changes induced by imbalance of glucose homeostasis and high intraglomerular pressure. Studies indicate that diseases are influenced not only by the environment but also by genetic factors. Among the genetic determinants, the polymorphisms in the MTHFR and VEGF genes stand out. These genes are oriented towards the synthesis of the enzyme methylenetetrahydrofolate reductase (MTHFR) and vascular endothelial growth factor (VEGF), respectively. The MTHFR enzyme plays an important role in the processing and processing cycle of methionine. VEGF is an important protein involved in signaling that stimulates vasculogenesis and angiogenesis. Thus, the objective of this study was to perform a molecular evaluation of MTHFR C677T and VEGF -141 A → C single nucleotide polymorphisms (SNPs) in diabetic patients with microvascular complications caused by hemodynamic and endothelial changes in a Brazilian population. A total of 345 were genotyped for polymorphisms using PCR-RFLP. The advanced RStúdio environment was used for statistical analysis. The logistic regression of the gene polymorphism in the MTHFR gene increases the risk (OR = 2.57, p = 0.003) of the diabetic individual developing ND. The polymorphism in VEGF gene reveals a 3.74-fold risk for DN (p = 0.001) in the eventual condition of diabetic patient. When analyzing the association of clinical variables and polymorphism in the MTHFR gene, the results showed no correlation. The association of clinical variables with the VEGF gene polymorphism, the results indicated the same in creatinine, in GFR and in diastolic blood pressure. Clinical associations observe a mechanism underlying the role of polymorphisms in renal dysfunction. Results suggest that polymorphisms MTHFR gene confer susceptibility to ND in the patients with DM.Item Avaliação do potencial in vitro e in vivo de tiossemicarbazida, tiossemicarbazida em nanopartícula de quitosana e tiossemicarbazida derivada do canfeno como antifúngico sobre Candida albicans(Universidade Federal de Goiás, 2016-05-06) Araújo, Deize Evangelista; Pereira, Maristela; http://buscatextual.cnpq.br/buscatextual/visualizacv.do?id=K4796262J0; Pereira, Maristela; Amaral, André Corrêa; Fernandes, Orionalda Fátima Lisboa; Ribeiro, Evandro leão; Borges, Clayton LuizCandidiasis is a serious health problem that affects a large number of people arround the world. The treatment of this disease is carried out with antifungals, which do not provide an effective cure, besides having resistance reports and high toxicity. In this context, it is necessary to study new antifungal compounds. In addition, the delivery form of the compounds in the living organism is also target of research, since the controlled release of compounds in biological systems can improve the mode of action of many drugs. Therefore, nanostructured systems, such as nanoparticles of chitosan, have been widely studied as controlled delivery of drugs. Then, this study aimed to evaluate the antifungal potential against Candida albicans of thiosemicarbazide (TSC), thiosemicarbazide-camphene (TSC-C) and chitosan nanoparticles incorporated with thiosemicarbazide (nanoTSC). In in vitro assays, the compounds were effective to inhibit nearly 100% of the fungus in concentrations of 1,37 mM to TSC, 0,02 mM to TSC-C and 0,27 mM to nanoTSC, and the most effective compound was TSC-C. NanoTSC and empty nanoparticles were also evaluated as to cell toxicity, and the concentration able to inhibit the fungus showed no cytotoxic activity. To confirm the values observed in in vitro assays, experiments were conducted using an animal model for vulvovaginal candidiasis. The results showed that TSC-C was more efficient in inhibiting C. albicans compared with TSC and nanoTSC, but was less effective than miconazole. However, histopathological analysis showed that the groups treated with the compounds under study, showed less intensity of damage to the vaginal epithelium and inflammatory infiltrates, compared with the positive control and the group treated with miconazole. Thus, the results suggest that TSC, nanoTSC and TSC-C are promising compounds in the treatment of vulvovaginal candidiasis.Item Efeitos de argentilactona sobre o perfil transcricional, parede celular e estresse oxidativo de Paracoccidioides spp(Universidade Federal de Goiás, 2014-06-02) Araújo, Felipe Souto; Pereira, Maristela; http://lattes.cnpq.br/1345781867765758; Pereira, Maristela; Schrank, Augusto; Cravo, Pedro Vitor Lemos; Soares, Célia Maria de Almeida; Rocha, Juliana ParenteParacoccidioides spp, dimorphic pathogenic fungi, is the etiologic agent of paracoccidioidomycosis (PCM). PCM is an endemic disease that affects at least 10 million people in Latin America, causing severe public health problem. The drugs used against pathogenic fungi have various side effects and have limited efficacy, therefore there is an inevitable and urgent medical needing for the development of new antifungal drugs. In the present study, we evaluated the transcriptional profile of Paracoccidioides spp exposed to argentilactone, a constituent of the essential oil of Hyptis ovalifolia. A total of 1,058 genes were identified, of which 208 were up-regulated and 850 down-regulated. The cell rescue, defense and virulence, with a total of 26 genes, was a functional category with a large number of genes induced, among them, heat shock protein 90 (hsp 90), cytochrome c peroxidase (ccp), hemoglobin ligant RBT5 (rbt5) and superoxide dismutase (sod). Quantitative real-time PCR revealed an increase in the expression level of all those genes. Enzymatic assay showed a significant increase in SOD activity. The reduced growth of Pbhsp90-aRNA, Pbccp-aRNA, Pbsod-aRNA, Pbrbt5-aRNA isolates in the presence of argentilactone indicates the importance of these genes in Paracoccidioides spp responding to argentilacone. Paracoccidioides spp cell wall was also evaluated. The results showed that argentilactone causes a decrease in the levels of polymers of the cell wall. These results suggest that argentilactone is a potential candidate for antifungal therapy.Item Análise de RNAs longos não codificantes do genoma de Arabidopsis thaliana (L.) Heynh(Universidade Federal de Goiás, 2017-03-07) Araújo, Vanessa Cristina da Silva; Novaes, Evandro; http://lattes.cnpq.br/0568272239145336; Novaes, Evandro; Vianello, Rosana Pereira; Coelho, Alexandre Siqueira GuedesLarge-scale sequencing of transcripts via RNA-Seq has been changing paradigms by demonstrating that transcription is prevalent throughout the eukaryotic genome. In these organisms, the vast majority of transcripts are non-coding (ncRNA). One type of RNA that has aroused great interest, given its prevalence, is long non-coding RNAs (lncRNAs), which are ncRNA with more than 200 nucleotides. However, little is known about the role and prevalence of these lncRNAs in plant genomes, even in model species such as Arabidopsis thaliana (L.) Heynh. The objective of this work was to identify lncRNAs in the Arabidopsis genome and to characterize their size, structure and nucleotide diversity. The sequences were obtained from previous work that sequenced total RNA from A. thaliana, grown under different light regimes, using Illumina Hiseq 2000 platform. These sequences were mapped into the reference genome with TopHat and assembled with Cufflinks. The assembled transcripts were compared with the genome annotation with Cuffcompare, to identify non-annotated transcripts. A total of 4,305 long putative RNAs were obtained, with 314 (7%) sense in relation to coding transcripts (mRNAs), 392 (9%) intergenic, 2,216 intronic (52%) and 1,383 (32%) antisense mRNAs. The lncRNAs obtained were filtered to eliminate those with coding potential, as well as those related to rRNA, tRNA and miRNA synthesis. A total of 3,710 high-confidence lncRNAs (HC-lncRNA) were obtained, of which 58.6% were not previously annotated. These HC-lncRNA emcompass a low proportion (~ 1%) lncRNAs in the genome of Arabidopsis thaliana. A functional enrichment analysis of Gene Ontology (GO) categories demonstrated that among genes containing lncRNAs there is a high proportion of categories linked to the localization and transport of proteins within the cell, as well as to nucleic acid binding. A gene expression analyses identified only 22 differentially expressed lncRNAs under the different light conditions in which samples were exposed. Using the SNP data from the 1001 genomes project, identified high nucleotide diversity within lncRNAs regions, indicating low conservation of the primary structure of these transcripts. The nucleotide diversity in regions of long noncoding RNAs is lower than in coding regions, but less than a diversity observed in neutral regions such as pseudogenes.Item Isolamento, caracterização e transferibilidade de marcadores microssatélites de Byrsonima cydoniifolia A. Juss. (Malpighiaceae)(Universidade Federal de Goiás, 2014-03-13) Bernardes, Vanessa; Telles, Mariana Pires de Campos; http://lattes.cnpq.br/4648436798023532; Telles, Mariana Pires de Campos; Vianello, Rosana; Borba, TerezaMolecular markers are identifiable DNA sequences found at specific sites in the genome and transmitted by the standard laws of inheritance from one generation to the next. Markers known as microsatellite are indicated for genetic population analysis; due they are widely distributed in the genome and their high polymorphic information content per locus. To support different management practices, genebanks, conservation studies is important to perform the analysis of genetic variability in natural populations of the species of interest. In the biome Cerrado there are inumerous medicinal and fruit trees that are used in a extractive way, among them are those of the gender Byrsonima sp. popularly known as murici, belonging to the family Malpighiaceae. In Brazil presents about 70 species with wide distribution in the Cerrado. The murici is a multi- potential plant and although not be domesticated, its economic exploitation can be viable. In this context, the objective was to develop a set of microsatellite markers for species Byrsonima cydoniifolia A. JUSS. and test the potential transferability of this set of markers for species Byrsonima crassifolia L. Kunth. The microsatellite regions were isolated using genomic libraries enriched for repetitive regions and then they were used for primer design. Using the primers designed temociclagem conditions were tested and the products were characterized using the DNA of 90 individuals of B. cydoniifolia and 24 indivuduals of B. crassifolia. Of these, 14 loci were polymorphic and 3 monomorphic in three populations of B. cydoniifolia. The number of alleles ranged from 3 to 17, with an average of 10.45 alleles per locus. The observed heterozygosity was similar to the expected heterozygosity, the values ranged from 0.20 to 0.92 and 0.43 to 0.92; respectively. 39 private alleles were identified. This set of markers showed high combined probability of paternity exclusion (Q) equal to 0.999 and combined probability of identity (I) equal to I=7.84x10-1. The global values for the fixation index (FIS) were not significant (P<0.05), whereas the values of FST=0,082 and FIT=0,143 were significant. For Byrsonima crassifolia species, nine pairs of primers produced good patterns of cross amplification in the study population, with a mean of 7.56 alleles per locus. The observed heterozygosity ranged from 0.250 to 0.958, which was higher than the expected heterozygosity ranging from 0.223 to 0.905. The combined probability of paternity exclusion (Q) was equal to 0.99847 and high combined probability of identity equal to 6.42x10-9. The strategy of development of microsatellite markers from the genomic library enriched methodology was efficient to generate a panel with eleven polymorphic microsatellite markers for B. cydoniifolia and nine polymorphic microsatellite markers successfully transferred to B. crassifolia, providing a useful tool for population genetic studies not only for the species B. cydoniifolia and B. crassifolia, but possibly for other related species evolutionarily.Item Genética geográfica de Hymenaea stigonocarpa (Fabaceae)(Universidade Federal de Goiás, 2015-09-29) Braga, Ramilla dos Santos; Telles, Mariana Pires de Campos; http://lattes.cnpq.br/4648436798023532; Nabout, João Carlos; Soares, Thannya NascimentoHymenaea stigonocarpa é uma espécie amplamente distribuída nas regiões do Cerrado brasileiro. Em função de diferenças morfológicas nas folhas, a literatura reconhece três variedades botânicas para esta espécie. Essas variedades são: H. stigonocarpa var. stigonocarpa, com folhas lisas; H. stigonocarpa var. pubescens, com folhas pilosas e H. stigonocarpa var. brevipetiolata, apresentando pecíolo curto, com folhas lisas ou pilosas. O conhecimento da divergência genética intervarietal e análise da estrutura genética espacial é interessante para entender o comportamento dessa espécie em subpopulações naturais e como os processos microevolutivos atuam na organização da sua variabilidade genética. Sendo assim, o objetivo deste trabalho foi verificar o nível de diferenciação genética existente entre as variedades botânicas, a magnitude da variabilidade genética entre e dentro as subpopulações de H. stigonocarpa, bem como o padrão espacial da variabilidade genética encontrada. Para tanto, foram realizadas expedições de coleta em regiões do Cerrado, onde amostrou tecido foliar de 32 subpopulações de H. stigonocarpa, num total de 1.034 indivíduos. A variabilidade genética foi acessada por meio de nove locos microssatélites, em que todas as subpopulações foram caracterizadas geneticamente por meio das estatísticas descritivas. A divergência genética entre variedades foi avaliada com a AMOVA, técnicas de agrupamento multivariadas e inferência Bayesiana. A estrutura genética foi estimada dentro e entre as 32 subpopulações de H. stigonocarpa. Estatísticas espaciais foram realizadas para avaliar a existência de padrões espaciais na variabilidade genética. Além disso, análises de complementariedade genética foram utilizadas para selecionar subpopulações potenciais para conservação. Os nove locos microssatélites foram polimórficos, apresentando níveis moderados de diversidade genética. A AMOVA revelou divergência genética considerável entre as variedades botânicas e aliada às análises de agrupamento indica que a variedade brevipetiolata é a mais diferente geneticamente quando comparado às variedades stigonocarpa e pubescens, que formaram um único grupo. As 32 subpopulações apresentaram baixa endogamia (f= 0,082; p<0,01), mas considerável diferenciação genética entre populações (FST= 0,161; p<0,01) e alta endogamia total (FIT= 0,230; p<0,01). Junto à estatística RST, que foi igual a 0,221 e significativa ao nível de 1% percebe-se que a mutação e deriva genética são importantes para a estruturação genética nesta espécie. As estatísticas espaciais foram obtidas para 30 subpopulações, pois a variedade brevipetiolata foi fortemente diferenciada sobre as demais e assim foi retirada destas análises. O teste de Mantel mostrou uma correlação matricial baixa, porém significativa (rm= 0,239; p=0,0014) entre distância geográfica e o FST par a par. O correlograma de Mantel identificou um padrão clinal na diferenciação genética, sugerindo que as subpopulações se comportam num modelo de isolamento por distância. A autocorrelação espacial, quantificada pelo índice I de Moran, foi baixa nas frequências alélicas entre os pares de subpopulações em cada classe de distância, gerando um correlograma médio sem padrão espacial nítido (rm=0,053; p=0,0005). O algoritmo de Monmonier identificou possíveis barreiras genéticas entre as subpopulações do extremo Norte do Cerrado, mas que precisam ser avaliadas com cautela. A complementariedade genética identificou 16 subpopulações que representam toda a variabilidade genética encontrada para a espécie nos locos avaliados e que foram selecionadas como prioritárias para conservação. As informações obtidas com este trabalho mostram que as estratégias reprodutivas da espécie são importantes para a conectividade genética entre subpopulações a longas distâncias, causando uma fraca estruturação genética espacial. Assim, é sugerido que não apenas o espaço esteja atuando na diferenciação genética, mas outros processos estocásticos. Além disso, a divergência genética entre variedades botânicas torna-se um componente importante para entender a sua estrutura genética, em que este trabalho traz um conhecimento relevante para estudos taxonômicos de H. stigonocarpa, podendo ser mais uma evidência para delimitação de espécies.Item Avaliação da susceptibilidade à mastite mediante análise de SNVs do promotor do gene CXCL8 em bovinos de aptidão leiteira(Universidade Federal de Goiás, 2019-03-12) Campos, Calebe Bertolino Marins de; Cruz, Alex Silva da; http://lattes.cnpq.br/5521953733966859; Cruz, Aparecido Divino da; http://lattes.cnpq.br/7868817504129985; Cruz, Aparecido Divino da; Silva, Daniela de Melo e; Amaral, Alliny das GraçasAgriculture is one of the sectors most responsible for Brazilian GDP, with 7.5% coming from livestock. Given the importance of livestock for the Brazilian economy, it is of great importance to study diseases that affect the national herds and their extrinsic and intrinsic factors. The objective of this study was to verify if there was association of clinical mastitis and subsequently of subclinical mastitis with alleles, genotypes and haplotypes found in six polymorphic sites of the promoter region of the CXCL8 gene. The animals from this study came from two herds of Goiás state, totalizing 98 animals and later the genomic sequences of interest in these animals were identified by qPCR using the KASP® kit (LGC Limited ®). In all the polymorphic sites three different genotypes were found and as in the distribution in the alleles and genotypes, four different haplotypes were identified. The alleles were found to be Hardy-Weinberg equilibrium and the loci were in linkage disequilibrium. The distribution of the animals of the animals in the positive and negative groups for clinical mastitis was different from the distribution found when the somatic cell count (SCC) of the milk was taken into account to separate the animals in the subclinical mastitis groups. No statistically significant associations were found between alleles, genotypes and haplotypes with the occurrence of clinical mastitis. However, there was a significant relationship between the GG genotype of the polymorphic site "KASP 4" (c-687) and the presence of subclinical mastitis, and the haplotype IL8-h2 had a protective effect against subclinical mastitis.Item Identificação de proteínas de superfície de Staphylococcus saprophyticus e análise de fatores de virulência(Universidade Federal de Goiás, 2014-06-09) Carvalho, Alex Jesus de; Rocha, Juliana Alves Parente; http://lattes.cnpq.br/7089231795367245; Rocha, Juliana Alves Parente; Coelho, Alexandre Siqueira Guedes; Baeza, Lilian CristianeThe Gram-positive bacterium Staphylococcus saprophyticus, one of the coagulasenegative staphylococci, is the second most common causative agent of urinary tract infection, affecting mainly sexually active women. Staphylococcus saprophyticus can cause acute diseases as pyelonephritis, sepsis, nephrolithiasis, endocarditis, urethritis, epididymitis and prostatitis. This work aims to identify Staphylococcus saprophyticus surface proteins by using a proteolytic shaving approach, a methodology that was established to identify surface-exposed protein domains by tripsinization of intact cells. The peptides obtained were treated by trypsin, reduced, alkylated and identified by nano-chromatography using a nanoACQUITY UPLCTM system (Waters) coupled to a SYNAPT Q-TOF mass spectrometer (Waters). The homology analysis was performed using the software ProteinLynx 2.3 (Waters). Through the shaving, it was possible to identify 219 proteins, many of them, described as virulence factors. Of total, 01 is cell wall protein, 09 are extracelular proteins, 19 are membrane proteins and 190 are citoplasmatic proteins. Besides of the lysis process, the presence of cytoplasmic proteins on cell surface can be due to the activity of export pathways not yet identified and many of these proteins can be proteins with moonlighting function, in other words, proteins that plays more of one function, it can, in this case, plays functions on S. saprophyticus cell surface related to bacterial virulence. The main proteins with moonlighting function include metabolic enzymes of the glycolytic pathway; enzymes of other metabolic pathways, such as, glyoxalate cycle; chaperones and proteins related with the proteic folding. The prediction of cellular localization was performed through LocateP database. The results of this research help to elucidate the strategies and machineries used by proteins during the adhesion, infection and proliferation, leading us to understand the interaction between the pathogenic bacteria S. saprophyticus and the human host. The knowledge about the proteins present on the cell surface is of extreme importance, because many of these proteins represent targets to new drugs, therapeutic antibodies or vaccines, since the pathogen cell surface is the first to contact with the host cells during the infection process.Item Desenvolvimento de marcadores microssatélites para Ololygon centralis (Anura: Hylidae) por sequenciamento de segunda geração com baixa cobertura(Universidade Federal de Goiás, 2018-03-12) Castro, Andrezza Arantes; Brito, Cíntia Pelegrinete Targueta de Azevedo; http://lattes.cnpq.br/9889075112366889; Telles, Mariana Pires de Campos; http://lattes.cnpq.br/4648436798023532; Telles, Mariana Pires de Campos; Brito, Cíntia Pelegrinete Targueta de Azevedo; Silva, Daniela de Melo e; Silva, Wilian VazThe ease access to Next Generation Sequence methodologies has improved the development of several projects, once it makes possible the de novo assembly of DNA sequences, creating a great amount of data and giving access to unknown genomes of different organisms. The anuran species Ololygon centralis belong to Hylidae family. It is considered as endemic of Cerrado Biome and little is known about its genomics. The aim of this work was to realize a draft of the genome of O. centralis in order to identify, to characterize, to quantify repetitive elements and to predict possible genes. Also, we aimed to develop microsatellite markers for the species. The genome draft assembled in 13989 scaffolds, which correspond to 4926069 bp with N50 = 336. The repetitive elements found in the O. centralis genome (1795 transposons, 957, microsatellite, one satellite DNA and 25 small nuclear RNAs) comprised 531337 bp. Between the retrotransposons, the LINE element (49,83%) was the most abundant, followed by LTR (48,66%) and SINE (1,56%). It was possible to identify coding genes for the 5S, 18S and 28S subunits of ribosomes. Also, 18 types of coding regions for tRNA and 26 putative genes were found in the O. centralis genome. From the microsatellite regions, it was possible to design 87 pairs of primers for the flanking sites. There were found 47 regions composed of tetranucleotides, 39 dinucleotides and on trinucleotide on the microsatellite sites. From them, 31 pairs of primers were selected for amplification and 18 showed good results. The annealing temperatures varied from 50° e 60° C. Seven markers showed polymorphism. From them, only 5 markers (PCE05, OCE11, PCE20, OCE21, OCE26) were used to characterize the genotype of 30 individuals. The medium number of alleles was 10, the allelic amplitude varied from 148 bp (OCE20) to 318 bp (OCE21). The Expected Heterozygosity was 0,7 and the observed was 0,5. The probability of parentage exclusion (Q) was 0,993 and the probability of combined identity (I) was 1,13x10-6. The global value of the fixation index (FST) was significant and equal to 0,2 (p<0.05). The data obtained from the NGS Illumina platform represent one important step for the knowledge of genomics of this species and of the anuran in general.Item Variabilidade genética de subpopulações de Mauritia flexuosa L.f (Arecaceae) em veredas do Estado de Goiás(Universidade Federal de Goiás, 2014-02-26) Corrêa, Kássia Marques; Chaves, Lázaro José; http://lattes.cnpq.br/9990967290797379; Soares, Thannya Nascimento; http://lattes.cnpq.br/5590256762396056Mauritia flexuosa is a large palm tree, commonly known as buriti that has wide distribution in the Cerrado, especially in association with fountains of waters and wetlands in formations known as palm swamps. The genetic variability of a species allows to elucidate the role of evolutionary processes in populations, contributing to the definition of efficient strategies for their use and conservation. Microsatellite markers are useful to quantify the level of variability among and within subpopulations. This study aimed to evaluate the genetic variability of six subpopulations of Mauritia flexuosa located in conserved and disturbed state of Goiás paths, based on microsatellite markers. 175 individuals were evaluated from six subpopulations. Genomic DNA was extracted from leaf tissue and subsequently quantified and diluted for performing polymerase chain reactions, using ten microsatellite loci. It was used an ABI3500 automatic sequencer, which acts through a capillary electrophoresis system, from which the genotypes were obtained. The array of genotypes obtained was used to estimate genetic diversity parameters among and within subpopulções. The analyzes were performed using GDA software. The average number of alleles per locus in subpopulations was 6,2, ranging from 5,6 to 7,0. The values of expected (He) and observed (Ho) heterozygosity were equal to 0,645 and 0,659, respectively, showing Ho value close to that expected by Hardy-Weinberg equilibrium. The analysis of population genetic structure revealed that subpopulations of M. flexuosa are moderately structured with θ = 0,099. The estimated value for f was not significant, as expected for dioecious species. The total fixation index considering all subpopulations was F = 0,08 , very close to the value of θ. This suggests that the genetic structure observed in these subpopulations is related to an effect of drifting associated with some restriction to gene flow between subpopulations. The limitation of gene flow between M. flexuosa subpopulations can be associated with the environment in which they occur, because the palm swamps have disjunct distributions and areas of dry land around them can act as barriers to gene flow. Another factor that could be related to this restriction to gene flow is the dispersion of seeds by zoochory, which becomes 13 difficult in fragmented environments and impracticable if it is kept in place unfavorable for seed germination. It was concluded that the sampled subpopulations showed a low level of inbreeding, and a relatively high genetic diversity. There is significant genetic structure, probably due to a slight isolation between subpopulations associated with the effects of genetic drift within subpopulations.Item Clonagem e expressão heteróloga, modelagem e interações intermoleculares da enolpiruvilchiquimato 3-fosfato sintase de Paracoccidioides brasiliensis(Universidade Federal de Goiás, 2017-08-07) Costa, Wanderson Lucas da; Pereira, Maristela; http://lattes.cnpq.br/1345781867765758; Pereira, Maristela; Lacerda, Elisângela de Paula Silveira; Neves, Bruno JuniorParacoccidioides spp. are thermodymorphic fungi that when inhaled by humans, these conidia find a favorable environment, changing to the yeast phase and becoming pathogenic causing paracoccidioidomycosis (PCM), one of the most prevalent systemic mycoses in Brazil. Some antifungals are used in the treatment of PCM. Treatment depends on the patient's progression and tolerability of each drug, but their treatment may be for long periods and cause various side effects in the patient. The chiquimate pathway is coordinated by 7 enzymes that perform consecutive steps to convert erythrose-4-phosphate and phosphoenol pyruvate (PEP) into chorismate. In microorganisms, this pathway is involved in the production of the amino acids phenylalanine, tyrosine and tryptophan; These amino acids are essential to the maintenance of these organisms. In this work, pGEX4T3 vector cloning and heterologous expression of Pb18 EPSP synthase belonging to the chiquimate pathway were performed. This protein was expressed in E. coli (DE3) strain and purified. Antibodies were produced for expression analysis of the protein in Western blot. The modeling of EPSP synthase was performed aiming to identify the amino acids involved in the active site. The pull down-GST assay with soluble Pb18 proteins allowed the identification of 40 proteins that interact with EPSP synthase. These proteins belong to different functional categories, which are involved with the availability of phosphoenol pyruvate, the substrate necessary for the functioning of the chiquimate pathway.Item Análise proteômica da fase leveduriforme do fungo patogênico Paracoccidioides sp durante a privação de nitrogênio(Universidade Federal de Goiás, 2014-03-13) Cruz-Leite, Vanessa Rafaela Milhomem; Borges, Clayton Luiz; http://lattes.cnpq.br/8867708267053410; Borges, Clayton Luiz; Bailão, Alexandre Melo; Brito, Wesley de Almeida; Casaletti, Luciana; Pereira, MaristelaThe fungi of the genus Paracoccidioides sp are etiologic agents of disease paracoccidioidomycosis (PCM), are termodimórficos and have the ability to move the saprobiótica form to yeast form at a temperature around 37 ° C. The uptake is essential for the growth and adaptation of the fungus in host tissue, where nitrogen-dependent pathways have close relationship with pathogenicity. Pathogenic organisms possess a regulatory system called Nitrogen Catabolic Represion which is induced when nitrogen availability is limiting in surround causing the expression of genes necessary for nutrient uptake when preferred sources such as glutamine and ammonia are scarce. This regulatory process comprises a complex system in the infectious process. This study aims to identify proteins regulated by nitrogen depletion, where the pathogenic fungus Pb01-like was analyzed as a proteomic response. Pb01-like was grown in minimal medium control MMcM (+N) and treated MMcM (-N) for 6 hours at 37 ° C, cytoplasmic proteins were extracted and subjected to tryptic digestion and quantification. The proteomic profile revealed an expression of 135 proteins, 40 proteins induced or identified the treaty, 58 repressed or identified control and 44 proteins were expressed constitutively. In silico analyzes showed that the gene is regulated by formamidase areA transcription factor that responds conditions of nitrogen depletion in Aspergillus nidulans and the surrounding medium the enzymatic activity for the gene of formamidase Pb01-like induced depletion of nitrogen during. The amino acid sequence of the regulatory gene areA important in the metabolism of nitrogen was aligned between Pb01-like, Pb18, Aspergillus nidulans and Neurospora crasssa where this alignment showed high identity and homology of conserved zinc finger domains and DUF 1752. Metabolic pathways such as fermentation, gluconeogenesis, protein synthesis, nitrogen metabolism were induced for the depletion of nitrogen, showing a possible modulation of the metabolism of the fungus in order to be able to adapt to environments with nitrogen limitation.Item Produção da enzima ciclodextrina glicosiltransferase por Bacillus sp. imobilizados em quitosana(Universidade Federal de Goiás, 2015-02-20) Es, Ismail; Amaral, André Corrêa; http://lattes.cnpq.br/8801299423520104; Amaral, André CorrêaProduction of a starch-degrading cyclodextrin glicosiltransferase (CGTase, EC 2.4.1.19) on solid-state culture and batch fermentation was evaluated by free and immobilized alkalophilic Bacillus sp on polymeric chitosan matrix. Immobilization procedure was performed by mixing bacterial cells with low molecular weight chitosan dissolved in HCl. Structure of free bacterial cell, polymeric chitosan matrix and immobilized cells were visualized by scanning electron microscopy (SEM). The images obtained from SEM showed that the procedure used for immobilization was easy, cheap and effective. Three different starch sources as substrate were used: potato, corn and cassava. Qualitative analysis of CGTase production was determined by colorless area formation on solid culture containing phenolphthalein. Enzymatic indices, which indicated the production of CGTase on solid culture, were calculated for both free and immobilized cells on different starch sources. Enzyme activity of CGTase was determined before and after each purification step: ammonium sulfate precipitation, starch adsorption and dialysis. Biomass growth and substrate consumption were analyzed by Flow cytometry and modified phenol-sulfuric acid assay, respectively. Free cells reached very high numbers (2.5 × 107 ml-1) during batch culture, besides; immobilized cells maintained initial inoculum concentration (2.5 × 105 ml-1) during enzyme production. The maximum enzyme activity achieved by free cells was 21.25 U (35 h), 14.65 U (40 h) and 19.16 U (33 h) on cassava, potato and cornstarch, respectively. During batch culture, immobilized cells produced CGTase with the enzyme activity of 24.375 U (16 h) for cassava, 24.375 U (9 h) for potato starch and 21.25 U (8.5 h) for cornstarch in a shorter cultivation time. Consequently, immobilization decreased the production time and increased enzyme activity of CGTase.Item Montagem e anotação funcional de sequências Gênicas de handroanthus impetiginosus (mart. ex DC) Mattos(Universidade Federal de Goiás, 2015-06-29) Fernandes, Lanusse Andrade; Novaes, Evandro; http://buscatextual.cnpq.br/buscatextual/visualizacv.do?id=K4704901A5Handroanthus impetiginosus, known as ipê roxo, is a species present in several Brazilian biomes, with ecological and economic importance. The species has suffered strong exploration thanks to the quality of its wood. Despite of its ecological importance, economic value and medicinal potential, there is virtually no genomic study published with this species. This work aimed to take advantage of the large sequencing capacity of next generation platforms to generate, assemble and functional annotate the sequences of the entire transcriptome of H. impetiginosus. Seeds were collected from two trees on the Campus II of the Universidade Federal de Goiás. The seeds were germinated, their aerial tissues collected and the RNA extracted and sent to the University of Illinois to be sequenced on the HiSeq 2500 platform (Illumina). A total of 148,359,530 sequencing reads were generated. The adapters and low-quality sequences were removed. Contaminating sequences were detected and eliminated, and the remaining sequences were normalized. Sequences were then assembled, annotated and polymorphisms were detected. A total of 190,885 scaffolds were assembled from 23,002 genes. Mimulus guttatus was the species with the most Blast best hit against the sequences of H. impetiginosus. Of all the Gene Ontology (GO) annotated sequences, the majority had categories for Biological Process (50.94%), followed by Cellular Component (28.34%) and Molecular Function (20.72%). The most frequent GO categories related to Biological Process were metabolic and cellular processes, while in Cell Components the most frequent were cell and organelles. Catalytic activity and binding of ions and molecules were the most frequent annotated categories of Molecular Function. As the sequences were generated from a pool of RNA from four plants, 25,169 InDels and 278,121 SNPs were identified. As expected, InDels are depleted in coding regions (CDS) when compared to SNPs. These results will be important for future studies with an interest in gene expression of the species. Furthermore, the possible SNPs identified can facilitate genetic studies and genomic H. impetiginosus populations.Item Análise proteômica de Paracoccidioides brasiliensis após estresse oxidativo(Universidade Federal de Goiás, 2017-03-28) Fernandes, Rosiane Rodrigues; Soares, Célia Maria de Almeida; http://buscatextual.cnpq.br/buscatextual/visualizacv.do?id=K4793575Z5; Borges, Clayton Luiz; Pigosso, Laurine LacerdaParacoccidioides brasiliensis is a thermodymorphic fungus, the etiological agent of Paracoccidioidomycosis, a systemic mycosis of high incidence in Latin America. During the infectious process, this fungus is subjected to oxidative stress, produced by host defense cells. Therefore, we sought to analyze the proteomic responses of the fungus when exposed to oxidative stress. Menadione is a compound that decreases the viability of yeasts of Paracoccidioides brasiliensis. The main results were the induction of antioxidant enzymes, such as superoxide dismutase, cytochrome c peroxidase, thioredoxin and peroxisomal catalase, enzymes that help the fungus to fight reactive oxygen species. In addition, proteins involved in metabolic pathways such as glycolysis, tricarboxylic acid cycle and glyoxylate cycle had increased expression, demonstrating a possible remodeling of fungus metabolism. There was the induction of enzymes of the pentose phosphate pathway, which has a prominent role in cells under oxidative stress, since it is an important mechanism producing NADPH (nicotinamide adenine dinucleotide phosphate), an electron donor relevant to counteract the deleterious effects of reactive species of oxygen Heat shock proteins were also upregulated during the oxidative stress to which the fungus was exposed. In this work we investigated the response of a member of the species Paracoccidioides brasiliensis, Pb339 to oxidative stress, including the adaptive mechanisms of this fungus to this condition caused in vitro or by the defense system promoted by the host.Item Aplicações toxicogenômicas em cultura de células expostas a nanomateriais e populações humanas expostas a pesticidas(Universidade Federal de Goiás, 2017-09-04) Franco, Fernanda Craveiro; Silva, Daniela de Melo e; http://buscacv.cnpq.br/buscacv/#/espelho?nro_id_cnpq_cp_s=9895211901348365; Parise, Michelle Rocha; Silva, Claudio Carlos da; Vieira, Thais Cidália; Costa, Emília Oliveira Alves; Silva, Daniela de Melo eThe interaction of some compounds and DNA molecule can cause genotoxic changes. The tests used to evaluate genotoxicity have evolved over the years, and they have great efficiency. Currently, several traditional methodologies are used, such as the comet assay, and some more modern, based on molecular biology. Considering the importance of detecting the effects caused by exposure to potentially genotoxic agents, this thesis presents three chapters with different approaches to this topic. The first chapter aimed at evaluating published studies on the mutagenic, genotoxic and oxidative effects caused by occupational exposure to pesticides. For this, a systematic review of articles published between the years of 2011 and 2015 was carried out, focusing on these exposure conditions. The results showed that 44% of the studies addressed only men and 44% approached men and women, and 78% used rural workers as an exposed population. The most evaluated class of pesticide was organophosphorus. The studies were separated by methodologies of analysis used and all showed some type of alteration in the populations occupationally exposed to pesticides. The second chapter, published in the journal Environmental Science and Pollution Research, considered the dengue scene in the state of Goias and techniques of mosquitoes control, and sought to understand the genotoxic effects caused by exposure of endemic agents to pesticides used against Aedes aegypti, being the first study published with this group of workers. The main pesticides used were insecticides and larvicides and an increase in DNA damage was observed in the exposed population, without influence of the lifestyle factors. In the whole transcriptome assay the differential expression of genes related to various chronic diseases was observed. Finally, the third chapter was carried out in collaboration with the Institute of Experimental Medicine of the Czech Academy of Sciences, aiming the determination of cytotoxic and genotoxic effects caused by in vitro exposure to different types of metallic nanomaterials. In nanomaterials that showed cell viability reduction (cytotoxic), an increase in genotoxicity was also identified. In addition, three of the four non-cytotoxic nanomaterials evaluated showed genotoxic effects. Studies aimed at detecting the compounds genotoxic potential are extremely important for the regulatory definition and identification of risks caused by exposure. In this study, the toxicological and genotoxic effects of different compounds and mixtures were evaluated using traditional and modern techniques, in vitro or by human biomonitoring, seeking to understand and determine the risks involved in exposure to these substances.