Made available in DSpace on 2015-03-26T13:42:31Z (GMT). No. of bitstreams: 1 texto completo.pdf: 483682 bytes, checksum: 46d5690ba1fa20fbb43c21ec1603c096 (MD5) Previous issue date: 2013-07-29 / Conselho Nacional de Desenvolvimento Científico e Tecnológico / Coffee breeding in Brazil promoted the development of cultivars with much higher productive potential, compared to traditional cultivars. Currently, 123 cultivars of Coffea arabica are registered in the National Register of Cultivars RNC/MAPA, of which eight are protected. Cultivars can only be registered if they are distinct, uniform and stable (DUS). Morphological descriptors have been generally used for DUS examination of a cultivar. However, commercial cultivars are increasingly coming phenotypically closer. This hinders a precise discrimination of these materials by such descriptors. Therefore, it is extremely important to develop an alternative to assist in DUS testing, because it will provide great benefits. The use of molecular markers can discriminate more accurately and safely cultivars candidates to registration and/or protection. Among the various types of molecular markers, microsatellites have been the most widely used because they are codominant, multi- allelic, highly polymorphic and locus-specific. This study aimed to establish a set of microsatellite markers for molecular characterization (fingerprinting) of coffee cultivars carrying rust resistance (Hemileia vastratrix Berk. Et Br). Thirty-four cultivars/progenies of C. arabica were assessed. For each cultivar/progeny, six plants were analyzed, which totaled 204 individuals. Thus, it was possible to verify the occurrence of genetic variability within and among the genetic materials evaluated. Thirty-one microsatellite primers were tested; 27 amplified, of which 20 were polymorphic. Based on the data of 16 polymorphic markers selected, a dendrogram was constructed, and a molecular profile of the cultivars was established in two distinct ways. The analysis of the dendrogram allowed distinguishing 29 of the 34 cultivars/progenies assessed. Fingerprinting analysis allowed defining 31 unique molecular profiles. / Os ganhos alcançados com o melhoramento genético do cafeeiro no Brasil resultaram na obtenção de cultivares com potencial produtivo muito superior ao das cultivares tradicionais. Atualmente, 123 cultivares de Coffea arabica estão registradas no Registro Nacional de Cultivares RNC/MAPA; destas, oito são protegidas. Para que uma cultivar possa ser registrada e ou protegida é obrigatório que ela seja distinta, homogênea e estável (DHE). Na comprovação de DHE de uma cultivar os descritores morfológicos tem sido os mais utilizados. No entanto, as cultivares comerciais são cada vez mais próximas fenotipicamente. Isso dificulta a discriminação precisa desses materiais por meio desses descritores. Assim sendo, uma alternativa que venha auxiliar nos testes de DHE é de extrema necessidade e proporcionará grandes benefícios. Nesse sentido, o uso de marcadores moleculares pode discriminar com maior precisão e segurança as cultivares as quais se deseja o registro e ou a proteção. Dentre os vários tipos de marcadores moleculares, os microssatélites tem sido os mais utilizados por serem codominantes, multialélicos, altamente polimórfico e loco específico. O presente trabalho objetivou estabelecer um conjunto de marcadores microssatélite para a caracterização molecular (fingerprinting) de cultivares de café portadoras de resistência à ferrugem (Hemileia vastatrix Berk. et Br.). Foram analisadas 34 cultivares/progênies de C. arabica. Para cada cultivar/progênie foram analisadas seis plantas, constituindo um total de 204 indivíduos. Dessa forma, pôde-se verificar a possível ocorrência de variabilidade genética entre e dentro dos materiais genéticos avaliados. Trinta e um primers microssatélites foram testados, 27 amplificaram, sendo 20 polimórficos. A partir dos dados de 16 marcadores polimórficos selecionados foi construído um dendrograma e de duas maneiras distintas, foi estabelecido o perfil molecular das cultivares. Pela análise do dendrograma foi possível distinguir 29 das 34 cultivares/progênies avaliadas. Pela análise de fingerprinting foi definido 31 perfis moleculares únicos.

Summary : The wild diploïd species Gossypium australe carry interesting agronomic characters such as resistance to wilt fusarium and "delay of the gossypol glands morphogenesis in the seed " that makes it an important source of variability for the genetic improvement of the main cultivated cotton species G. hirsutum. One of the approach to introgress these characters is to isolate and exploit monosomic alien addition lines (MAAL). In order to isolate new MAAL of G. australe in G. hirsutum, the [2(G.hirsutum x G.australe)x G.hirsutum] pentaploid was backcrossed as male parent to G. hirsutum. Among the 253 BC1 derivatives obtained, 106 plants (42%) presented morphological alterations attributed to presence of G. australe chromatin. To define an SSR linkage group for each of the 13 G. australe chromosomes, 42 plants representative of the phenotypic variability observed in the BC1 generation and seven alien addition lines already isolated in our laboratory were analyzed using SSR markers developed from the G. hirsutum species. Out of the 150 SSR markers used, 100 % amplified G. australe DNA and 84 (56 %) generated 89 polymorphic loci. All these loci but two have been assigned, by means of an cluster algorithm, to 13 linkage groups assumed to match up to the 13 chromosomes of the diploid species. On this basis, about 60% of the analyzed plants were multisomic addition lines, 20%, MAAL while 20 % carrying no markers were supposed to be euploid. The newly isolated MAAL appeared to be the same as those already available. Five disomic alien addition plants carrying at least one additional chromosome different from the chromosomes of G. australe previously isolated in a monosomic addition configuration were selfed and the BC1S1 progenies obtained have been analyzed with SSR markers and GISH. Five new MAAL of G. australe in G. hirsutum have thus been isolated. In order to monitor the potentialities of using MAAL for the transfer of genetic material from the additional chromosome to the genetic background, the transmission frequency and integrity of the supernumerary chromosome have been analyzed with SSR markers in the self-progeny of five MAAL. Three of them revealed a transmission frequency significantly lower than the 3:1 expected ratio, one MAAL presented an exclusive preferential transmission of the additional chromosome. In these four MAAL the alien chromosome was transmitted almost unaltered. With the fifth MAAL the alien chromosome was normally transmitted but was altered in half of the plants containing G. australe chromatin. One of the investigated MAAL characterized by its brown fiber produced few plants carrying also white fibers. It has been shown that this mosaicism was due to the loss of the alien supernumerary chromosome. The complete loss of this chromosome seems to be linked to its fragmentation. Résumé : L'espèce diploïde sauvage Gossypium australe possède des caractères agronomiques d'intérêt tels que la résistance au fusarium et le "retard à la morphogenèse des glandes à gossypol" qui en font une importante source de variabilité pour l'amélioration génétique de la principale espèce de cotonnier cultivé G. hirsutum. Une des approches pour l'introgression de ces caractères est la production et l'exploitation de lignées monosomiques d'addition (LMA). Pour isoler les LMA de G. australe sur G. hirsutum, le pentaploïde [2(G.hirsutum x G.australe)x G.hirsutum] a été rétrocroisé comme parent mâle avec l'espèce tétraploïde. Sur les 253 graines obtenues, 106 (42%) ont donné des plantes présentant une morphologie nettement distincte de celle de G. hirsutum. Cette différence a été attribuée à la présence de chromosomes de G. australe. Afin de définir des groupes de liaison pour chacun des chromosomes de G. australe, 42 plantes représentatives de la variabilité phénotypique observée ainsi que 7 lignées d'addition déjà isolées ont été sélectionnées et analysées avec des marqueurs SSR développés sur l'espèce tétraploïde. Tous les 150 marqueurs utilisés ont amplifié l'ADN de G. australe et 84 (56%) ont généré 89 loci polymorphes. Tous ces loci, sauf deux, ont pu être assignés, par classification numérique, à 13 groupes de liaison supposés correspondre aux 13 chromosomes de l'espèce diploïde. Sur cette base, 60% des plantes analysées sont des plurisomiques d'addition; 20%, des LMA tandis que 20 % ne portant aucun marqueur ont été supposées euploïdes. Les nouvelles LMA isolées s'étant révélées être identiques à celles déjà isolées, 5 plantes disomiques d'addition portant au moins un chromosome non-encore isolé à l'état monosomique d'addition ont été autofécondées et leur descendance analysée avec des marqueurs SSR et par la GISH. Cinq nouvelles LMA ont pu ainsi être isolées. Afin d'étudier les potentialités d'utilisation de la méthode des LMA pour le transfert de matériel génétique de l'espèce sauvage vers l'espèce cultivée, la fréquence de transmission et l'intégrité du chromosome surnuméraire, a été analysée avec des marqueurs SSR dans une génération autofécondée de cinq LMA. Trois lignées ont donné un taux de transmission inférieur au ratio attendu de 3:1, chez la quatrième lignée le chromosome surnuméraire a été transmis à toute la descendance. Pour ces quatre lignées le chromosome additionnel a été transmis presque inaltéré. Avec la cinquième lignée, le chromosome additionnel a été transmis suivant le taux attendu mais a été altéré dans la moitié des plantes contenant de la chromatine de G. australe. Une des lignées analysées caractérisée par la couleur brune de ses fibres a produit quelques plantes portant également des fibres blanches. Il a été montré que ce mosaïcisme de la couleur des fibres était dû à la perte du chromosome additionnel. Cette perte semble être liée à une fragmentation du chromosome.

ABSTRACT Triticum sphaerococcum (AABBDD, 2n = 6x = 42) is a land race of wheat known from the Indian subcontinent. It has several favourable characters including short and strong culms, hemispherical grains with a shallow crease (that may increase the yield of white flour), higher protein content compared to bread wheat (T. aestivum), and resistance to drought, and yellow rust caused by Puccinia striiformis. However, an unfavourable characteristic of T. sphaerococcum is its lower yield compared to bread wheat. Being a land race, the sphaerococcum wheat is poorly studied. This study was undertaken to increase knowledge of the physiology and genetics of this land race and determine if it may be possible to separate the favourable characters of T. sphaerococcum from its unfavourable characters. Plant height in bread wheat is controlled by many genes. ‘Reduced Height’ (Rht) genes which differ in their response to externally applied gibberellic acid (GA3) are responsible for the short stature of modern bread wheat varieties. Therefore, GA3 was used to probe the relationship between the semidwarf sphaerococcum phenotype and the Rht gene. T. sphaerococcum variety Sp5 showed a unique “seedling response” to externally applied GA3 when compared with T. aestivum varieties harbouring Rht1, Rht2, Rht8, Rht12, Rht13 or Rht18 alleles. A mapping population of doubled haploids was generated through wide hybridisation of F1 (Sp5 x Otane) with Zea mays. A genome-wide scan of Sp5 and Otane (parents) using 348 microsatellite (SSR) markers showed that only 169 of these markers (49%) were polymorphic between the parents. A DArT profiling yielded 348 markers that were polymorphic between the parents. Microsatellite markers and DArT markers were used to create a genetic map. The mapping population was phenotyped and a quantitative trait loci (QTL) analysis was performed for component traits of the complex sphaerococcum trait including plant height, spike length, awn length, yield, grain shape and crease size. Results of the QTL analysis indicated that it may be difficult to separate the favourable characters of T. sphaerococcum from its unfavourable characters through mutation because the component traits of the complex sphaerococcum trait may be under pleiotropic control of the Sp gene. The hypothesis that T. sphaerococcum originated through a mutation in T. aestivum was tested through induced mutation using gamma rays. Mutants from sphaerococcum-type to aestivum-type were isolated and phenotyped. Sphaerococcum-type mutants also were isolated and characterised from mutated aestivum-type wheat suggesting a possible origin of T. sphaerococcum through a mutation in T. aestivum.

ABSTRACT Triticum sphaerococcum (AABBDD, 2n = 6x = 42) is a land race of wheat known from the Indian subcontinent. It has several favourable characters including short and strong culms, hemispherical grains with a shallow crease (that may increase the yield of white flour), higher protein content compared to bread wheat (T. aestivum), and resistance to drought, and yellow rust caused by Puccinia striiformis. However, an unfavourable characteristic of T. sphaerococcum is its lower yield compared to bread wheat. Being a land race, the sphaerococcum wheat is poorly studied. This study was undertaken to increase knowledge of the physiology and genetics of this land race and determine if it may be possible to separate the favourable characters of T. sphaerococcum from its unfavourable characters. Plant height in bread wheat is controlled by many genes. ‘Reduced Height’ (Rht) genes which differ in their response to externally applied gibberellic acid (GA3) are responsible for the short stature of modern bread wheat varieties. Therefore, GA3 was used to probe the relationship between the semidwarf sphaerococcum phenotype and the Rht gene. T. sphaerococcum variety Sp5 showed a unique “seedling response” to externally applied GA3 when compared with T. aestivum varieties harbouring Rht1, Rht2, Rht8, Rht12, Rht13 or Rht18 alleles. A mapping population of doubled haploids was generated through wide hybridisation of F1 (Sp5 x Otane) with Zea mays. A genome-wide scan of Sp5 and Otane (parents) using 348 microsatellite (SSR) markers showed that only 169 of these markers (49%) were polymorphic between the parents. A DArT profiling yielded 348 markers that were polymorphic between the parents. Microsatellite markers and DArT markers were used to create a genetic map. The mapping population was phenotyped and a quantitative trait loci (QTL) analysis was performed for component traits of the complex sphaerococcum trait including plant height, spike length, awn length, yield, grain shape and crease size. Results of the QTL analysis indicated that it may be difficult to separate the favourable characters of T. sphaerococcum from its unfavourable characters through mutation because the component traits of the complex sphaerococcum trait may be under pleiotropic control of the Sp gene. The hypothesis that T. sphaerococcum originated through a mutation in T. aestivum was tested through induced mutation using gamma rays. Mutants from sphaerococcum-type to aestivum-type were isolated and phenotyped. Sphaerococcum-type mutants also were isolated and characterised from mutated aestivum-type wheat suggesting a possible origin of T. sphaerococcum through a mutation in T. aestivum.

Made available in DSpace on 2014-07-29T14:52:05Z (GMT). No. of bitstreams: 1 claudio takao karia.pdf: 2242096 bytes, checksum: 8b502fdb3ff4995409e0345270eaaed3 (MD5) Previous issue date: 2008-08-21 / Plant genetic resources maintained ex situ, in germplasm banks, are few used in agricultural systems and genetic plant breeding programs. One of the main reasons of this incipient use is the difficulty to obtain information about stored accessions, especially the characterization and preliminary evaluation data. Stylosanthes guianensis is a predominantly self-pollinated and diploid species, which has a great number of this stored accessions and potential for use in agricultural systems. There are approximately one thousand accessions of the S. guianensis in Embrapa‟s germplasm bank. Moreover, only one genotype of this species is now available to commercial growers in Brazil, the cultivar Mineirão. To promote the use of these resources, this study aimed to characterize accessions of S. guianensis, stored in the germplasm bank of Embrapa, using morpho-agronomic traits and microstellites markers (SSR). In morpho-agronomic characterization 535 accessions were evaluated using 23 quantitative traits. The data were analyzed by a principal component analysis (PCA), the Ward's agglomerative hierarchical clustering method, and by univariate analysis of variance, associated to a Tukey test for comparisons among means of the established groups. Thirteen similarity accession groups were established, five of them were considered the ones with the highest potential use in agricultural systems. Regarding these potential groups, it is possible to reduce from 535 to 126 accessions to be thoroughly evaluated. The molecular characterization was made in 437 accessions, using seven microsatellites fluorescent primers, and the detection of the DNA fragments generated by PCR (Polimerase Chain Reaction) was accomplished by a capillary electrophoresis. The seven loci provided 45 alleles (6.43 alleles/locus), of which four were private alleles. Both, principal coordinates analysis and cluster analysis, in this case, by UPGMA agglomerative method, showed a tendency to group accessions together by botanical varieties (canescens, guianensis and pauciflora), except for the microcephala variety. The correlation between the genetic and morpho-agronomic dissimilarities was highly significant, however, it was of low magnitude (r = 0.23, P ≤ 0.0001). This indicates that both evaluations are important and should be performed to achieve a more complete characterization of these accessions and, possibly, of the species. Taking account the germplasm accessions with agronomic potential, selected based on the morpho-agronomic evaluation, several of them presented significant genetic dissimilarities to the Mineirão cultivar. These accessions should be better assessed for agronomic traits, and crossed with this cultivar in breeding programs of the species. The results of this research will allow increasing the efficiency and effectiveness of the germplasm bank activities, helping the choice of genotypes to be tested in agronomic evaluations, and stimulating further researches with these genetic resources. / Os recursos genéticos vegetais armazenados ex situ, em geral, são pouco utilizados nos sistemas de produção e no melhoramento genético de plantas. Um dos principais motivos deste uso incipiente é a dificuldade de se obterem as informações sobre os acessos armazenados, sobretudo os dados de caracterização e avaliação preliminar. A espécie Stylosanthes guianensis, diplóide e preferencialmente autógama, possui grande quantidade desses acessos e potencial de utilização em sistemas agrícolas. Somente no Banco de Germoplasma da Embrapa existe uma coleção com cerca de mil acessos registrados de S. guianensis. Por outro lado, apenas um genótipo dessa espécie é hoje comercializado no Brasil, a cultivar Mineirão. Para promover a utilização desses recursos, o presente trabalho teve como objetivo caracterizar os acessos de S. guianensis, armazenados no Banco de Germoplasma da Embrapa, utilizando-se caracteres morfoagronômicos e marcadores de locos microssatélites (SSR). Na caracterização morfoagronômica, foram avaliados 535 acessos acerca de 23 caracteres quantitativos. Os dados correspondentes foram estatisticamente investigados empregando-se a análise de componentes principais, a análise de agrupamento, pelo método aglomerativo de Ward, e a análise de variância (univariada), associada ao teste de Tukey, para comparar as médias dos grupos identificados. Foram estabelecidos treze grupos de acessos similares, dos quais, cinco foram considerados de maior potencial para a utilização nos sistemas agrícolas. Considerando-se apenas os grupos potenciais, é possível reduzir o número de acessos a serem avaliados mais detalhadamente, de 535 para 126 acessos. Na caracterização genético-molecular foram considerados 437 acessos da coleção, utilizando-se sete primers microssatélites, marcados com fluorescência, e a detecção dos fragmentos de DNA obtidos via PCR (reação da polimerase em cadeia) foi feita por meio da técnica de eletroforese em capilar. Os sete locos produziram 45 alelos (6,43 alelos/loco), dos quais quatro foram alelos privados. Tanto na análise de coordenadas principais, como na análise de agrupamento, neste caso pelo método UPGMA, os acessos demonstraram tendência de se reunirem em grupos, conforme as variedades botânicas da espécie (canescens, guianensis e pauciflora), exceto para a variedade microcephala. A correlação entre as dissimilaridades genéticas e morfoagronômicas foi altamente significativa, porém, de baixa magnitude (r = 0,23; P ≤ 0,0001), indicando que ambas as avaliações são importantes e devem ser realizadas para a melhor caracterização desses acessos e, possivelmente, da espécie. Considerando-se os grupos com potencial agronômico, selecionados na avaliação morfoagronômica, identificaram-se acessos geneticamente bastante dissimilares da cultivar Mineirão, que devem ser mais bem avaliados agronomicamente, para compor possíveis cruzamentos com essa cultivar, em programas de melhoramento da espécie. Assim, os resultados deste trabalho permitirão aumentar a eficiência e a eficácia das atividades no Banco de Germoplasma em estudo, subsidiar a escolha de genótipos a serem testados em avaliações agronômicas, podendo, ainda, estimular outras pesquisas com esses recursos genéticos.