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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 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 Montagem e análise do transcritoma do mogno africano (Khaya grandifoliola C. DC.)(Universidade Federal de Goiás, 2019-07-25) Soares, Sabrina Delgado; Novaes, Evandro; http://lattes.cnpq.br/0568272239145336; Novaes, Evandro; Telles, Mariana Pires de Campos; Soares, Thannya Nascimento; Coelho, Alexandre Siqueira Guedes; Vianello, Rosana PereiraKhaya grandifoliola C. DC. is a tropical species with high value wood in international markets. Despite the high interest for reforestation and timber production, virtually nothing is known about the genetic and genomic aspects of the species. With the development of next-generation sequencing platforms (NGS), the characterization of transcriptomes can be performed on a large scale and at affordable cost. In this context, the objective of the work was to discover K. grandifoliola genes by using RNA-Seq technique and to identify those differentially expressed between the leaf and the xylem K. grandifoliola tissues. Total RNA was extracted from leaf and xylem samples of four trees planted in the experimental area of the UFG. mRNA samples were sequenced on the Illumina HiSeq 2000 platform, in paired-end mode 2 × 100 bp. Sequencing resulted in a total of 857 million sequences which, after quality control, decreased to 808 million with an average size of 24 to 85 bp. Trinity program was used to reassemble the transcriptome, obtaining 116,289 transcripts for a total of 36,271 genes. Transcripts ranged from 524 to 12,031 bp with a mean of 1,366 bp and N50 of 1,657 bp. Analyses of gene expression identified 13,626 differentially expressed genes (DEGs) (FDR <0.05) between the two tissues. Of these, 5,438 had higher expression in leaves and 8,188 in xylem. A functional enrichment analysis of Gene Ontology categories among the DEGs was performed by the Blast2GO program. This analysis identified 26 GO terms enriched among the most active DEGs in xylem. Among genes up-regulated in leaves, 48 enriched terms were found. As expected, the categories identified as enriched between DGEs point to biological processes and cellular components specific of each tissue. For example, among the most active genes in leaf tissue, terms associated with thylakoids (GO:0009579), plastids (GO:0009536) and photosynthesis (GO:0015979) were identified. On the other hand, for the xylem, terms related to carbohydrate metabolism (GO:0005975), cell wall (GO:0071554) and cytoskeletal proteins (GO:0008092) were significantly enriched. To the best of our knowledge, this is the first report of large-scale gene sequences for K. grandifoliola. Differential expression analysis allowed us to identify which genes are most active in leaf and xylem. Analysis in iPath3 identified the metabolic routes activated in leaf and xylem tissues. A BlastX search against the orange proteome (Citrus sinensis) revealed the presence of 95 DEGs annotated for the enzymes of the limonoid, a secondary metabolite with medicinal properties.Item Diversidade genética em população de melhoramento de mogno africano (Khaya ivorensis A. Chev.)(Universidade Federal de Goiás, 2014-08-04) Soares, Sabrina Delgado; Novaes, Evandro; http://lattes.cnpq.br/0568272239145336The strong demand for hardwood drives selective logging and degradation of forests ecossystems in Brazil. Due to its wood value and predatory logging, Brazilian mahogany (Swietenia macrophylla) is at risk of extinction. Currently, its trees are protected and banned from logging by the Brazilian Institute of Environment (IBAMA). Monoculture of the Brazilian mahogany is proven unsuccessful due to the attack of the Hypsipyla grandella larvae. An alternative to Brazilian mahogany plantation is the African mahogany (Khaya ivorensis), a species with similar high valued wood properties. The Universidade Federal de Goiás in partnership with Mudas Nobres Company started a breeding program with Khaya ivorensis. The objective of this study was to evaluate the genetic diversity and divergence among 53 superior trees selected and cloned as part of the breeding strategies. Clones were selected from three different populations, planted in two farms in Pará state and originating from Ivory Coast and Tanzania populations. Twelve seedlings of Khaya senegalensis were also used as a control group in the analyses of genetic divergence. The individuals were genotyped with eight microsatellite loci developed for K. senegalensis, by capillary electrophoresis on the ABI-3100 (Applied Biosystems) platform. The software GeneMapper (Applied Biosystems) was used to genotype the alleles. The average number of alleles per locus was 5.875. The average expected heterozygosity was lower (HE = 0.563) than the average observed heterozygosity (HO = 0.738). Therefore, the average intrapopulation fixation index was negative (f = - 0.314) indicating that high inbreeding depression is decreasing the frequency of homozygous when compared to what would be expected under Hardy-Weinberg Equilibrium. The average fixation index between populations was estimated at θ = 0.008, indicating that only 0.8% of the genetic variability is due to differences between populations. The dendrogram generated from the matrix of Rogers’ genetic distance showed two distinct groups, separating the control group with K. senegalensis from the K. ivorensis selected trees. The group composed of K. ivorensis had nodes with weak bootstrap consistency indicating weak genetic structure among selected trees. The lack of genetic structure was confirmed by a Bayesian approach on the Structure (version 2.3.4) program. The genetic diversity observed within the selected breeding population is comparable to that of natural populations of African and Brazilian mahoganies. The genetic distance estimated with this work will guide the selection of divergent progenitors to be crossed.Item Expressão gênica diferencial de quatro espécies da Aliança Tabebuia em resposta ao deficit hídrico(Universidade Federal de Goiás, 2017-03-10) Sobreiro, Mariane Brom; Novaes, Evandro; http://buscatextual.cnpq.br/buscatextual/visualizacv.do?id=K4704901A5; Coelho , Alexandro Siqueira Guedes; Brondani, Claudio; Novaes, EvandroConsidering the rate of increase in average annual temperature and the seasonality of rainfall in several regions of the country, investigations on the mechanisms of plant’s response to low water availability become relevant. Tabebuia Alliance species - monophyletic clade of the Bignoniaceae family are commonly known as ipe – are distribuited in areas with different soil and climatic conditions. This feature makes them an interesting model to understand mechanisms tolerance’s to abiotic stresses. For each species there were two groups: control group, which had maintained irrigation; low water availability group, which irrigation was stopped and the experiment continued until the subtrate reached 40% of field capacity. The main objective of this work was to identify differentially expressed genes (DEG) in four species - two from the Brazilian savannah (T. aurea and Handroanthus ochraceus) and two from seasonally dry forests (H. impetiginosus and H. serratifolius). Then, RNA was extracted from the plants for sequencing on the Illumina platform with paired-end sequences of 100 base pairs (bp). Sequences were evaluated for quality control and mapped onto the genome of H. impetiginosus to identify DEG using the R software. The DGEs obtained by DESeq2 were subjected to functional enrichment analysis and potential changes in the level of expression of genes encoding enzymes of particular metabolic pathways. In all tools, H. serratifolius species showed the highest number of DGE (4908 noDESeq2), while H. ochraceus had the lowest number of DGE (6 in DESeq2). Functional enrichment analyzes demonstrated that the species presented, individually or collectively, typical responses of low water availability such as decrease of photosynthetic rate, increase of proline and increase of starch degradation. Although species share some responses, the complexity of organisms does not allow them to exhibit identical behaviors.Item Genômica populacional de Handroanthus serratifolius e Tabebuia aurea(Universidade Federal de Goiás, 2018-03-09) Vieira, Lucas Donizetti; Novaes, Evandro; http://lattes.cnpq.br/0568272239145336; Collevatti, Rosane Garcia; Silva, Daniela de Melo e; Lima, Natácia Evangelista dePopulation genomics studies in Neotropical species are scarce, however, they are of great importance for the understanding of the neutral and adaptive variations, in order to develop strategies of management and conservation for the species. In this context our objective was to verify the role of natural selection in the distribution of populations diversity in Handroanthus serratifolius and Tabebuia aurea. Here, we study population genomics based in sequence capture using 24,751 pre-developed probes. For H serratifolius were sampled c. 64,000 SNPs in 144 individuals from 16 populations. For T aurea were sampled c. 18,000 SNPs in 48 individuals from 6 populations. Genetic differentiation was high (FST = 0.194, p <0.001) in H. serratifolius and intermediate (FST = 0.144, p <0.001) in T. aurea. The inbreeding coefficient was low in both species, however, in three populations of H. serratifolius it was intermediate or high. The genetic structure of populations presented two genetic groups for the populations sampled in the two species. H. serratifolius with high genetic differentiation and T. aurea with admixture in all populations. We did not find SNPs outliers in the distribution of FST values, due to the wide distribution of values found for the populations of the two species. In the search for correlation between environmental variables and changes in allelic frequency H. serratifolius presented only three SNPs, inside six genes and T. aurea presented 16 SNPs, inside 25 genes. The functional annotation of these genes showed relationship with molecular processes, transposable elements and one in T. aurea related to resistance to diseases. In conclusion, our data show a selection signal in H. serratifolius for only three loci with local adaptation signal. In T. aurea our data show a greater number of locos (16 locos) with local adaptation signal.