Doctor of Philosophy / Genetics Interdepartmental Program / Jesse A. Poland / Wheat is perhaps the most important component in human diet introduced since the conception of modern agriculture, which provides about 20% of the daily protein and calorie intake to billions of people. Adaptable to wide range of climates, wheat is grown worldwide, lending it the potential to mitigate the imminent risk of food security for future population of 9.5 billion people. For developing improved crop varieties in the future, genetic diversity is a key factor in plant breeding. Constraints in wheat evolution and artificial selection practices have resulted in erosion of this ingredient in elite germplasm. However, wheat wild relatives, such as Ae. tauschii, D-genome donor of wheat, are a storehouse for unexploited genetic diversity that can be used for improving wheat for disease and insect resistance, yield, quality, and tolerance to abiotic stresses. More than 1700 genebanks around the world hold over 7 million accessions of these wild relatives. These genebanks are expensive to maintain, therefore, efficient curation is necessary. We developed and implemented a protocol to identify duplicate accessions using genomic tools. Implementing this approach with three genebanks, we identified over 50% duplicated accessions across genebanks. There are over a million Triticeae accessions held collectively, and it is likely as more number of genebanks are tested, there will be decreasing number of unique accessions. Selecting and utilizing the wild genetic diversity is no easy task. Historically, breeders and geneticists have chosen the accessions primarily based on associated phenotypic data. Unless focusing on a targeted trait, this practice is imperfect in capturing the genetic diversity with some other limitations, such as confounding phenotypic data with the testing environment. Utilizing next-generation sequencing methods, we selected a MiniCore consisting of only 40 accessions out of 574 capturing more than 95% of the allelic diversity. This MiniCore will facilitate the use of genetic diversity present in Ae. tauschii for wheat improvement including resistance to leaf rust, stem rust, Hessian fly, and tolerance to abiotic stresses. Hessian fly is an important insect pest of wheat worldwide. Out of 34 known resistance genes, only six have been mapped on the D sub-genome. With swift HF evolution, we need to rapidly map and deploy the resistance genes. Some of the undefeated HF resistance genes, such as H26 and H32, were introgressed from Ae. tauschii. In this study, we mapped three previously known genes, and a new gene from Ae. tauschii accession KU2147. Genes were mapped on chromosomes 6B, 3D, and 6D. Further, identification and cloning of resistance genes will enhance our understanding about its function and mode of action. In conclusion, wild wheat relatives are genetically diverse species, and utilizing the novel genetic diversity in Ae. tauschii will be fruitful for wheat improvement in the wake of climate change to ensure future food security to expected 2 billion newcomers by 2050.

Orientadores: Anete Pereira de Souza, Cacilda Borges do Valle / Tese (doutorado) - Universidade Estadual de Campinas, Instituto de Biologia / Made available in DSpace on 2018-08-13T19:56:51Z (GMT). No. of bitstreams: 1 Cancado_LeticiaJungmann_D.pdf: 2473309 bytes, checksum: ab9e0a498f29fb6c76b2ae1bb9583bc5 (MD5) Previous issue date: 2009 / Resumo: Brachiaria é um gênero de gramíneas composto por cerca de 100 espécies. Nas últimas décadas, algumas destas espécies vêm sendo amplamente usadas como forrageiras nas regiões tropicais. Uma importante coleção de germoplasma, que preserva 443 acessos e amostra 14 espécies diferentes, foi introduzida no Brasil na década de 1980 e está estabelecida na Embrapa Gado de Corte, Campo Grande/MS. Esta coleção vem sendo usada no Programa de Melhoramento Genético de Forrageiras Tropicais da Empresa Brasileira de Pesquisa Agropecuária. São cinco as espécies de maior valor para este programa de melhoramento: B. brizantha, B. decumbens, B. dictyoneura, B. humidicola e B. ruziziensis. Este trabalho visou ao desenvolvimento de marcadores moleculares do tipo microssatélites para B. brizantha e B. humidicola e a caracterização da diversidade genética existente em acessos e cultivares destas cinco espécies. Foi desenvolvido um total de 28 microssatélites polimórficos para B. brizantha e 27 para B. humidicola. Vinte microssatélites desenvolvidos para B. brizantha foram usados na caracterização da diversidade genética intra-específica em 160 acessos e cinco cultivares desta espécie. Dentre estes 20 locos, sete foram usados para caracterizar a variabilidade interespecífica entre estes 165 genótipos de B. brizantha e 13 acessos de B. decumbens, sete de B. dictyoneura, 42 de B. humidicola e 16 de B. ruziziensis. Os 27 microssatélites desenvolvidos para B. humidicola foram usados na caracterização da diversidade genética existente nos 58 acessos desta espécie, conservados nesta coleção, além de duas cultivares. A diversidade interespecífica acessada através de similaridades genéticas mostrou que os marcadores utilizados foram capazes de distinguir as cinco espécies estudadas, sendo que B. brizantha, B. decumbens e B. ruziziensis revelaram-se mais similares entre si e mais dissimilares em relação a B. dictyoneura e B. humidicola. Apesar disso, uma análise Bayesiana revelou que B. brizantha e B. decumbens compartilham pools alélicos entre si, não compartilhados pelas outras espécies. Do mesmo modo, B. dictyoneura e B. ruziziensis compartilham um pool gênico entre si, enquanto que B. humidicola apresenta um pool gênico distinto. Estes resultados combinados estão discutidos. A análise de diversidade intraespecífica em B. brizantha mostrou que os genótipos avaliados foram posicionados em três grupos principais de diversidade, sendo que esta diversidade não está fortemente estruturada. A análise intraespecífica em B. humidicola mostrou uma clara distinção do único acesso sexual desta coleção em relação aos demais, todos apomíticos, e revelou que a diversidade genética nesta coleção está mais bem estruturada que a encontrada em B. brizantha. x / Abstract: Brachiaria is a genus that comprises about 100 species. In the last decades, some of its species have been widely used as forages in the Tropics. A germplasm collection with 14 species represented by 443 accessions was introduced to Brazil in the decade of 1980 and is maintained at Embrapa Beef Cattle, Campo Grande, central Brazil. This collection has been used in the Tropical Forages Breeding Program of the Brazilian Agricultural Research Corporation. The most valuable species, considering agronomical aspects, are: B. brizantha, B. decumbens, B. dictyoneura, B. humidicola and B. ruziziensis. The main goal of this work was the development of microsatellite markers for B. brizantha and B. humidicola and the characterization of the genetic diversity found within these species in both inter- and intraspecific approaches. A total of 28 polymorphic microsatellites is described for B. brizantha, while for B. humidicola we present 27 polymorphic loci. Twenty microsatellites of B. brizantha were used for assessing the genetic variability in 160 accessions and five cultivars of this species. Out of these twenty loci, seven were used to evaluate the 165 genotypes of B. brizantha and 13 accessions of B. decumbens, 7 of B. dictyoneura, 42 of B. humidicola and 16 of B ruziziensis. Twenty seven loci reported for B. humidicola were used in the intraspecific characterization of the genetic diversity of the whole collection of this species (58 accessions) and two cultivars. The interspecific genetic diversity revealed through similarities showed that the markers used were able to distinguish the five species studied. B. brizantha, B. decumbens and B. ruziziensis were the most similar, while B. humidicola and B. dictyoneura were closer to each other. Besides, a Bayesian analysis showed that B. brizantha and B. decumbens share exclusive allelic pools not present in the other species. Likewise, B. dictyoneura e B. ruziziensis share another allelic pool, while B. humidicola did not share genic pools with any other species. The intraspecific genetic diversity survey in B. brizantha revealed that genotypes were positioned within three major groups, but the genetic variability does not seem to be very structured. The intraspecific survey in B. humidicola showed a clear distinction between the one sexual accession of this species and all the other apomictic accessions, and unveiled that the genetic diversity within this species is more strongly structured than in B. brizantha. / Doutorado / Genetica Vegetal e Melhoramento / Doutor em Genetica e Biologia Molecular

Submitted by Ricardo Cedraz Duque Moliterno (ricardo.moliterno@uefs.br) on 2016-01-26T21:56:33Z No. of bitstreams: 1 TESE_Anderson_FINAL.pdf: 3427530 bytes, checksum: 027d12f66415d71dc327e860609665fe (MD5) / Made available in DSpace on 2016-01-26T21:56:33Z (GMT). No. of bitstreams: 1 TESE_Anderson_FINAL.pdf: 3427530 bytes, checksum: 027d12f66415d71dc327e860609665fe (MD5) Previous issue date: 2015-06-29 / Conselho Nacional de Pesquisa e Desenvolvimento Cient?fico e Tecnol?gico - CNPq / Coordena??o de Aperfei?oamento de Pessoal de N?vel Superior - CAPES / Funda??o de Amparo ? Pesquisa do Estado da Bahia - FAPEB / Eplingiella fruticosa (Salzm. Ex Benth.) Harley & JFB Pastore is an aromatic species, native, occurring in six states in northeastern Brazil (Bahia, Sergipe, Pernambuco, Paraiba, Rio Grande do Norte and Cear?). Popularly known as "alecrim de vaqueiro", is commonly found in street markets of the region and used to combat pain and seizures. Reviews in mice and in vitro studies show analgesic activity, vasodilating, cardioprotetiva, anti-inflammatory and larvicidal of its essential oil and of different types of the leaves extract. Recent studies show great variability in essential oil chemical composition of E. fruticosa, related to soil and climatic conditions and different plant organs. Thus, the species has great potential for exploration both agronomic, and by pharmaceutical companies. The aim of this study was to evaluate the vegetative propagation capacity and characterize previously Eplingiella genotypes, through morphological, agronomic, phytochemicals and molecular data. In Chapter I, two experiments were conducted: the first tested the effect of three substrates and the second evaluated five concentrations of IBA and three periods of cultivation. The design was a randomized block design with four replications. We evaluated survival percentage (% S), percentage of rooted cuttings (% EE), root length (CRE), number of shoots (NBE), dry mass of leaves (MSF), root dry weight (MSR) and total dry matter (MST). In Chapter II, twelve genotypes were collected, propagated vegetatively and transplanted. Twelve months after transplantation were assessed 12 quantitative traits, eight morphological and agronomic four. In Chapter III, the total DNA was extracted, then 20 primers were tested, of which nine were selected because they have better electrophoretic profiles agarose gel (2%). The binary matrix was computed in GEOCOMPAR II. It is estimated the diversity of the genetic structure parameters and the data were subjected to Bayesian analysis, and Neighbor-joining dendrogram and principal component analysis (PCA) based on matrix of Nei distances. And in Chapter IV, samples of 100g of leaves each repetition per genotype were used in the essential oil hydrodistillation in Clevenger type apparatus for three hours, quantifying the content. The identification of the compounds and their contents was performed by GC (FID) and GC / MS data 15 and the major compounds were used in diversity analysis. They have been made to cluster analysis and canonical variables, using as dissimilarity measure the Mahalanobis distance (D2). In the first experiment of Chapter I, significant differences were found for CRE, NBE, MSF, MSR and MST, with the best performance for the commercial substrate. In the second, positive effects have been identified both the addition of AIB as the cultivation time on the CRE variables, NBE, MSF and MSR, reaching maximum increment to the estimated concentration of 1.5 g L-1, at 60 days of cultivation. In Chapter II, there was significant variation by F test (p <0.01) for the CF features, LF, CBD, CBE, LP, and MFF MSF. The genotypes formed two groups for almost all variables, by Scott-Knott test (p <005), except for LP, which formed three. The EF002 and EF003 genotypes presented the highest levels for almost all variables. There was the formation of three groups for both UPGMA and for the canonical variables (CV). The characteristics that most contributed to the formation of groups were CBE, MFF and CF. The genotypes EF002, EF003, EF005 and EF012 stood out because they have higher genetic distances. In CHAPTER III, primers produced 131 polymorphic bands. The diversity index of Nei (Ne) ranged between 0.31 and 0.39, while Shannon (I) ranged between 0.33 and 0.48. The percentage coefficient of genetic differentiation (Gst) was 0.29. In AMOVA most of the variation was within populations (69%), while among populations was 27% and 4% among species, indicating a good genetic structure. The average value of Fst was 0.175, demonstrating intermediate differentiation between populations. The structure of the Bayesian analysis method revealed three possibilities for the formation of groups (K = 2; = 6; 8 =;), however, it presented many migrants and high level of mixing individuals. The dendrogram generated by the Neighbor-Joining method confirmed the formation of two groups, with good support for major clades (100%). PCA analysis in the first two axis explained 21.06% of the total variation among populations. Finally, in Chapter IV, the genotypes were classified into four clusters: 1 - EF001 genotypes, EF006, EF007, EF008, EF010, EF011 and EF012 with E-caryophyllene and bicyclogermacrene as major; 2 - EF002 and EF003 genotypes, with the majority same as the previous group, however, percentage with average about 30% higher; 3 - EF004 and EF005 genotypes that showed a greater production of E-caryophyllene; and 4 - with EF009 genotype, forming a single group to present ?-pinene as balanced majority and percentage among the rest. This result was confirmed by canonical variables, which explained 76% of the variation. The bicyclogermacrene compounds, 1,8-cineol, ?-copaene and spathulenol represented the most important variables for analysis. / Eplingiella fruticosa (Salzm. ex Benth.) Harley & J.F.B. Pastore ? uma esp?cie arom?tica, nativa, que ocorre em seis estados do nordeste brasileiro (Bahia, Sergipe, Pernambuco, Para?ba, Rio Grande do Norte e Cear?). Popularmente conhecida como ?alecrim de vaqueiro?, ? comumente encontrada em feiras livres da regi?o e utilizada no combate a dores e convuls?es. Avalia??es em camundongos e in vitro comprovam atividades analg?sicas, vasodilatadora, cardioprotetiva, antinflamat?ria e larvicida do seu ?leo essencial e de diferentes tipos de extrato de suas folhas. Estudos recentes apontam grande variabilidade na composi??o qu?mica do ?leo essencial de E. fruticosa, relacionada ?s condi??es edafoclim?ticas e aos diferentes ?rg?os vegetais. Sendo assim, a esp?cie apresenta grande pot?ncial de explora??o tanto agron?mica, quanto por ind?strias farmac?uticas. O objetivo geral deste estudo foi avaliar a capacidade de propaga??o vegetativa e caracterizar, previamente, gen?tipos de E. fruticosa, por meio de dados morfol?gicos, agron?micos, fitoqu?micos e moleculares. No CAP?TULO I, foram conduzidos dois experimentos: o primeiro testou o efeito de tr?s substratos e o segundo avaliou cinco concentra??es de AIB e tr?s per?odos de cultivo. O delineamento foi em blocos casualizado, com quatro repeti??es. Avaliou-se percentagem de sobreviv?ncia (%S), percentagem de estacas enraizadas (%EE), comprimento da raiz (CRE), n?mero de brota??es (NBE), massa seca de folhas (MSF), massa seca de raiz (MSR) e massa seca total (MST). No CAP?TULO II, doze gen?tipos foram coletados, propagados vegetativamente e transplantados. Doze meses ap?s o transplante foram avaliadas 12 caracter?sticas quantitativas, sendo oito morfol?gicase quatro agron?micas. No CAP?TULO III, o DNA total foi extra?do, em seguida 20 iniciadores foram testados, dos quais nove foram selecionados por apresentarem melhores perfis eletrofor?ticos em gel de agarose (2%). A matriz bin?ria foi computada no GEOCOMPAR II. Estimou-se os par?metros de diversidadee a estrutura gen?tica os dados foram submetidos ? an?lise Bayesiana, al?m de dendrograma Neighbor-joining e an?lise de componentes principais (PCA) com base na matriz de dist?ncias de Nei. E no CAP?TULO IV, amostras de 100g de folhas de cada repeti??o por gen?tipo foram utilizadas na hidrodestila??o do ?leo essencial, em aparelho tipo clevenger, durante tr?s horas, quantificando-se o teor. A identifica??o dos compostos e seus teores foi realizada por CG (DIC) e CG/EM e os dados de 15 compostos majorit?rios foram utilizados nas an?lises de diversidade. Foram procedidas an?lise de agrupamento e de vari?veis can?nicas, utilizando como medida de dissimilaridade a dist?ncia generalizada de Mahalanobis (D2).No primeiro experimento do CAP?TULO I, foram verificadas diferen?as significativas para CRE, NBE, MSF, MSR e MST, com melhor desempenho para o substrato comercial. No segundo, foram identificados efeitos positivos tanto da adi??o de AIB quanto dos tempos de cultivo sobre as vari?veis CRE, NBE, MSF e MSR, atingindo incremento m?ximo com a concentra??o estimada de 1,5 g L-1, aos 60 dias de cultivo. No CAP?TULO II, houve varia??o significativa, pelo teste de F (p<0,01), para as caracter?sticas CF, LF, CBD, CBE, LP, MFF e MSF. Os gen?tipos formaram dois grupos para quase todas as vari?veis, pelo teste de Scott-Knott (p<005), exceto para LP, que formou tr?s. Os gen?tipos EF002 e EF003 apresentaram as maiores m?dias para quase todas vari?veis. Houve a forma??o de tr?s grupos, tanto para UPGMA quanto para as vari?veis can?nicas (VC). As caracter?sticas que mais contribu?ram para a forma??o dos grupos foram CBE, MFF e CF. Os gen?tipos EF002, EF003, EF005 e EF012 se destacaram por apresentarem maiores dist?ncias gen?ticas. No CAP?TULO III, os iniciadores produziram 131 bandas polim?rficas. O ?ndice de diversidade de Nei (Ne) variou entre 0,31 e 0,39, enquanto Shannon (I) variou entre 0,33 e 0,48. O percentual do coeficiente de diferencia??o gen?tica (Gst) foi de 0,29. Na AMOVA a maior parte da varia??o ficou dentro das popula??es (69%), enquanto entre popula??es foi de 27% e entre esp?cies de 4%, indicando uma boa estrutura??o gen?tica. O valor m?dio de Fst foi 0,175, demonstrando diferencia??o intermedi?ria entre as popula??es. As an?lises de estrutura pelo m?todo Bayesiano revelou tr?s possibilidades de forma??o de grupos (K=2;=6;=8;), no entanto, apresentou muitos indiv?duos migrantes e elevado n?vel de miscigena??o. O dendograma gerado pelo m?todo de Neighbor-Joining confirmou a forma??o de dois grupos, com boa sustenta??o para os principais clados (100%). Na an?lise de PCA os dois primeiros axis explicaram 21,06% da varia??o total entre as popula??es. Por fim, no CAP?TULO IV, os gen?tipos foram classificados em quatro clusters: 1 - gen?tipos EF001, EF006, EF007, EF008, EF010, EF011 e EF012, com E-cariofileno e biciclogermacreno como majorit?rios; 2 - gen?tipos EF002 e EF003, com os mesmos majorit?rios que o grupo anterior, no entanto, com percentuais m?dios cerca de 30% superiores; 3 - gen?tipos EF004 e EF005, que evidenciaram uma maior produ??o de E-cariofileno; e 4 - com gen?tipo EF009, formando um grupo isolado por apresentar ?-pineno como majorit?rio e percentuais equilibrados entre os demais. Esse resultado foi confirmado pelas Vari?veis Can?nicas, que explicou 76% da varia??o. Os compostos biciclogermacreno, 1,8-cineol, ?-copaeno e espatulenol representaram as vari?veis de maior import?ncia para a an?lise.