Hybridisation between introduced and endemic species causes conservation concerns, but also provides us with an opportunity to study the dynamics of gene flow between two species as they first meet. Japanese sika deer (Cervus nippon) were introduced to the British Isles at a number of locations at the beginning of the 20th century. In the intervening time, sika have spread and their range now extends across approximately 40% of Scotland, where they overlap with that of native red deer (C. elaphus), with which they hybridise. In this study we focus on the consequences of one particular introduction that took place at Carradale, on the Kintyre Peninsula in 1893. First, I assessed the current state of hybridisation using a sample of 735 red and sika deer samples collected in 2006/7 from forestry blocks throughout the Kintyre Peninsula. Genetic analysis was conducted with a panel of 22 highly differentiated microsatellite loci and one mtDNA marker. Population admixture analysis of the microsatellite data was conducted with the Bayesian clustering programme STRUCTURE. Over most of the study area, levels of introgression into red and sika deer were low and were consistent with a scenario of very occasional F1 hybridisation followed by backcrossing. There was, however, one forestry block where 43% of individuals could be defined as hybrids. Second, I developed a branching process model of introgression via backcrossing, to assess whether variation in introgression across microsatellite loci could be interpreted as a signature of selection, or could in fact be attributed to stochastic processes. If only a few hybridisation events have contributed to the hybridising population, the pattern of introgression, even with a large number of genetic markers, will be highly stochastic. This pattern of neutral variation in introgression can have high enough variance that it could be mistaken for selection. Therefore, even if strong selection is acting, it may not be possible to distinguish its effects from neutral variation. Third, I analysed trends in hybridisation and introgression over 15 years on the peninsula, through analysis of a dataset of 1513 red and sika deer samples at 20 microsatellite and a mtDNA marker. There was little evidence of change in the extent of hybridisation and introgression over time. MtDNA introgression was predominantly from red deer into sika. Recent introgression into sika on the peninsula can be explained by a very small number of F1 hybridisation events (~10) via analysis of the number of alleles that have introgressed from polymorphic red deer into the genetically homogenous sika population (a similar analysis cannot be conducted for introgression into red deer). Finally, I conducted a regression analysis of genetic hybrid scores against phenotypic traits to assess the effect of hybridisation on phenotype. Hybridisation has caused changes in the weight of sika-like deer and red-like females. Hybridisation has caused changes in incisor arcade breadth of both populations and jaw length (a proxy for skeletal size) in sika-like females. However, there is no evidence that hybridisation has caused changes in kidney fat (a measure of condition) or pregnancy rates in either population. In conclusion, even a small number of F1 hybridisation events can lead to extensive introgression and the timing and spatial distribution of these events is likely to have a large impact on the structure of a recently hybridising population - stochastic factors dominate both the distribution of hybrid individuals and the distribution of the genes that introgress following a hybridisation event. In red deer and sika deer, increasing phenotypic similarities of the two populations caused by hybridisation are likely to facilitate further breakdown between the two species. It is possible that breakdown in assortative mating between the two species could occur across their range.

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Doctor of Philosophy / Genetics Interdepartmental Program - Plant Pathology / Bikram Gill / The diploid D genome species, Aegilops tauschii Coss. (2n=2x=14,DD) has provided numerous genes for resistance to diseases and insect pests that attack bread wheat (Triticum aestivum L. (2n=6x=42, AABBDD). Wheat production is currently threatened by broadly virulent races of the ‘Ug99’ lineage of wheat stem rust caused by the fungus, Puccinia graminis f.sp. tritici Pers. & Eriks. Screening of a large set of Ae. tauschii germplasm for resistance to TTKSK (Ug99) identified potentially novel sources of resistance. To expedite utilization of TTKSK resistance from Ae. tauschii, a direct hybridization approach was established that integrates gene transfer, mapping and introgression into one process. Direct crossing of Ae. tauschii accessions with an elite wheat breeding was used to initiate transfer of resistance. Genetic mapping of resistance was accomplished during gene transfer through development of BC[subscript]2 mapping populations. Bulked segregant analysis of BC[subscript]2F[subscript]1 genotypes at 70 SSR loci across the D genome identified the chromosome locations of stem rust resistance genes and facilitated genetic mapping. Using this approach, TTKSK resistance from CDL4424 and TA1662 was mapped on chromosome arm 1DS flanked distally by Xwmc432 and proximally by Xwmc222 at 4.4 cM, TA10187 on 6DS linked to Xcfd49 at 1.9 cM and TA10171 on 7DS linked Xwmc827 at 0.9 cM. TTKSK resistance from additional Ae. tauschii accessions CDL4366, TA1615, TA1642, TA1693 and TA1718 has been recovered in segregating populations but await mapping. Altogether, TTKSK resistance from eight Ae. tauschii accessions has been introgressed to a hard winter wheat genetic background. Three new stem resistance genes have been tagged with molecular markers for marker assisted breeding and will provide valuable material for stem rust resistance breeding and gene pyramids for effective control of stem rust.

Plant domestication led to a loss of genetic variation in many crops, due to the excessive emphasis in the selection of edible organs (root, leaf, stem or fruit) and the low selection pressure for other traits in the cultivated environment. This \'genetic erosion\' led to loss of alleles associated with resistance to environmental stresses, such as drought and salinity, which can in turn culminate in productivity losses. In tomato (Solanum lycopersicum L.), it is possible to tap into a reservoir of valuable genetic variation in its wild relatives. Identification of genetic variants associated with tomato domestication, and with stress resistance mechanisms which may have been lost during domestication, could be used to aid in breeding programs. In the present work, which was divided into two chapters, we carried out crosses between the wild species S. pennellii and the miniature tomato cultivar Micro-Tom (MT) and created two introgression lines (ILs), one with reduced organ size and another with increased drought tolerance. In the first chapter, we report the characterization and mapping of the IL denominated as Tiny organs and reduced yield (Toy). Toy harbors a S. pennellii genome segment on chromosome 7 and presents a considerable reduction in both vegetative (leaves) and reproductive (fruit) organs. We discuss how this could be a relevant trait underpinning tomato domestication. In the second chapter, we describe the drought tolerance mechanism of the IL Water Economy Locus in Lycopersicon (Well). Well harbors a S. pennellii genome segment on chromosome 1 and shows lower hydraulic conductance, possibly related to decreased xylem vessel size. The results shown suggest that this lower hydraulic conductance promotes a disturbance in the soil-plant-atmosphere hydraulic continuum leading to changes in stomatal behavior, which, in turn, are probably related to the delayed wilting of Well under conditions of water deficit. / A domesticação das plantas levou a uma perda de variação genética em muitas culturas, devido à ênfase excessiva na seleção de órgãos comestíveis (raiz, folha, caule ou fruto) e a baixa pressão de seleção para outras características no ambiente cultivado. Essa \"erosão genética\" levou à perda de alelos associados à resistência de diversos estresses ambientais, como seca e salinidade, os quais, por sua vez, podem conduzir a perdas significativas na produtividade das plantas. Entretanto, no tomate (Solanum lycopersicum L.), é possível acessar um banco valioso de variação genética nas espécies selvagens relacionadas. Assim, a identificação de variantes genéticas associadas ao processo de domesticação do tomateiro e a mecanismos de resistência a estresses ambientais, os quais podem ter sido perdidos durante a domesticação, pode auxiliar em programas de melhoramento do tomateiro e de outras culturas de interesse comercial. Diante disso, no presente trabalho, o qual foi dividido em dois capítulos, realizamos cruzamentos entre a espécie selvagem, S. pennellii, e a cultivar miniatura de tomateiro Micro-Tom (MT) para criamos duas linhas de introgressão (ILs), uma com tamanho de órgão reduzido e outra com maior tolerância à seca. No primeiro capítulo, relatamos a caracterização e mapeamento da IL denominada como Tiny organs and reduced yield (Toy). O genótipo Toy carrega um segmento do genoma de S. pennellii no cromossomo 7 e apresenta uma considerável redução em órgãos vegetativos (folhas) e reprodutivos (frutos). Os resultados obtidos conduziram a uma discussão de como esse genótipo pode ser relevante para a domesticação do tomateiro, devido ao seu impacto no tamanho de diversos orgão. Por outro lado, no segundo capítulo, descrevemos o mecanismo de tolerância à seca da IL Water Economy Locus em Lycopersicon (Well). Plantas Well carregam um segmento do genoma de S. pennellii no cromossomo 1 e exibem uma menor condutância hidráulica, possivelmente relacionada ao tamanho reduzido do vaso xilemático. A menor condutância hidráulica do genótipo Well conduz a perturbações no contínuo solo/planta/atmosfera levando a mudança no comportamento estomático, que, por sua vez, provavelmente está relacionado a maior resistência ao murchamento apresentada por esse material em condições de déficit hídrico.

The study of natural genetic variations affecting organogenic capacity in tomato (Solanum lycopersicum) is attractive due to the existence of several tomato wild relatives with enhanced organogenic capacity. The characterization of such variations is relevant not only in order to manipulate plant development, but also to understand its ecological and evolutionary significance. The objective of this work was to characterize three tomato loci whose alleles from the wild relative S. pennellii enhance in vitro shoot and root regeneration, and analyze their involvement in the acquisition of competence phase. In the first manuscript, we report the genetic and physiological characterization of the loci Rg3C, Rg7H and Rg8F. The S. pennellii alleles were introgressed into the tomato genetic model cv. Micro-Tom (MT), creating the near isogenic lines (NILs) MT-Rg3C, MT-Rg7H and MT-Rg8F. In the second manuscript we present a comparative analysis between the Near-Isogenic Lines (NILs) MT-Rg3C and MT-Rg1. Since Rg1 was proposed to be a key gene in the acquisition of competence, and was mapped in the chromosome three, it is believed that Rg3C is probably equivalent to the Rg1 allele from S. peruvianum. After the introgression of the loci into the MT background, the NILs presented enhanced regeneration of both roots and shoots, confirming that the loci were successfully introgressed. The analysis of the time for acquisition of competence and induction, together with the molecular characterization of the NILs, indicate that the genes present in the loci Rg3C, Rg7H and Rg8F affect in vitro regeneration by distinct pathways. While Rg3C decreased the time required for both acquisition of competence and induction, the other loci seem to influence only the time of acquisition of competence, in the case of Rg8F, or the time of induction, in the case of Rg7H. Additionally, although MT-Rg3C has an enhanced shoot branching phenotype, MT-Rg7H and MT-Rg8F did not differ from MT in this trait. This indicates that enhanced in vitro shoot formation in tomato is not necessarily related to a deleterious high branching phenotype. Comparative analyses of MT-Rg1 and MT-Rg3C strongly indicate that Rg1 and Rg3C are alleles of a same gene controlling regeneration capacity. Integrating Rg1 and Rg3C mapping information, we were able to narrow the number of candidate genes for Rg1/Rg3C to only 27, which were also analyzed and discussed. / O estudo de variações genéticas naturais afetando a capacidade de organogênese in vitro em tomateiro (Solanum lycopersicum) é promissor devido a existência de uma série de espécies selvagens relacionadas ao tomateiro, que apresentam alta capacidade organogênica in vitro. A caracterização de tais variações é relevante não apenas com o objetivo de manipulação do desenvolvimento vegetal, mas também com o intuito de entender o significado ecológico e evolutivo de tal característica. O objetivo desse trabalho foi caracterizar três loci de tomateiro, cujos alelos vindos de seu parente selvagem S. pennellii aumentam a capacidade de regeneração de gemas caulinares e radiculares in vitro, e analisar o envolvimento de tais loci na fase de aquisição de competência para regeneração. Nós apresentamos no primeiro capítulo a caracterização genética e fisiológica dos loci Rg3C, Rg7H e Rg8F. Os alelos de S. pennellii foram introgredidos na cultivar modelo Micro-Tom (MT), criando as linhagens quase isogênicas (Near Isogenic Lines - NILs) MT-Rg3C, MT-Rg7H e MT-Rg8F. No segundo capítulo nós analisamos comparativamente as NILs MT-Rg3C e MT-Rg1. Uma vez que Rg1 foi proposto como gene chave na aquisição de competência, e assim como Rg3C está localizado no cromossomo 3, acredita-se que Rg3C seja provavelmente ortólogo ao gene Rg1 de S. peruvianum. Após a introgressão dos loci na cultivar MT, as NILs, assim como esperado, apresentaram alta taxa de regeneração tanto de gemas caulinares, quanto de radiculares in vitro, confirmando que os loci foram devidamente introgredidos. A análise do tempo de aquisição de competência e indução, juntamente com a caracterização molecular das NILs, indicam que os genes localizados nos loci Rg3C, Rg7H e Rg8F afetam a regeneração in vitro através de rotas distintas. Enquanto Rg3C diminui o tempo necessário tanto para a aquisição de competência quanto para indução de gemas caulinares, os outros dois loci parecem influenciar apenas a aquisição de competência, no caso de Rg8F, ou a indução de gemas caulinares, no caso de Rg7H. Além disso, apesar de MT-Rg3C apresentar alta ramificação, MT-Rg7H e MT-Rg8F não diferiram de MT nesse aspecto, o que evidencia que a formação de gemas caulinares in vitro não está necessariamente relacionada ao aumento da ramificação. As análises comparativas entre MT-Rg3C e MT-Rg1 indicam fortemente que Rg1 e Rg3C sejam dois alelos de um mesmo gene controlando a alta capacidade de regeneração. Através do cruzamento dos dados de mapeamento disponíveis para esses dois alelos foi possível diminuir o número de genes candidatos à Rg1/Rg3C para apenas 27 genes, que são apresentados nesse trabalho.

Currently, the genus Thraupis Boie, 1826 is a monophyletic group with seven species, all of which have high molecular and morphological support. Nevertheless, the phylogenetic relationship of the species still unclear: T. abbas is the sister species of T. ornataT. palmarum clade, and a second group within the genus is composed by the T. episcopusT. sayaca clade. Furthermore, in the remaining species group, one of the species, T. glaucocolpa, has not been included in any of the previous molecular studies, even it was believed to be close related with T. sayaca. Moreover, the last species within the genus, T. cyanoptera, has an uncertain position in the genus phylogenetic tree. The T. episcopusT. sayacaT. galucocolpa species complex includes 18 subspecies and a high morphological variation and a wide distribution which includes overlapping zones of T. episcopus and T. sayaca, makes taxa identification almost impossible. Nonetheless, previous molecular studies had only used samples from two individuals of T. episcopus and one of T. sayaca. Furthermore, the group does not have taxonomic stability, as shown by the multiple changes, which occur at different levels: moving from one genus to another or from species to subspecies level etc. To check the genus, I analyze 1171 specimens. The morphometric analysis outcomes show the weight as the most variable and important measure and T. cyanoptera as the only clearly different species within taxonomic units. Finally, I did a phylogenetic analysis based on two mitochondrial genes (Cyt- and ND2), in addition to three nuclear introns (intron 3 of MUSK gen, intron 5 of TGFB2 gen and a piece of the intron 5 of the BF5 gen). I performed the extractions from tissues collected at different localities around the natural distribution of the species, with emphasis on T. episcopus and T. sayaca. I ran independent locus RAxML analysis and haplotypes networks and used to group the samples on genetic taxonomic units. RAxML and haplotypes analysis shows a close relationship between T. episcopus and T. sayaca with high probably introgression process within. Furthermore, exposed a genetic structure within T. episcopus. I used this genetic taxonomic units to ran a multilocus species tree with a calibrated molecular clock. The species tree suggests that the origin of the genus Thraupis was between 5.5 and 7.5 million years before present, in the Messinian age. Also recovers T. glaucocolpa is the oldest linage in the genus and shows a relation between the morphological traits with the genetic structure within T. episcopus. Finally, I suggest synonymizing several subspecies and elevating to species level the subspecies T. episcopus cana, based on the morphological and molecular data. / O gênero Thraupis Boie, 1826 é um grupo monofilético composto por sete espécies. Entretanto, as relações entre estas espécies continuam obscuras. Thraupis abbas é a espécie irmã do clado T. ornataT. palmarum. Um segundo grupo é composto pelo clado T. episcopusT. sayaca. Por outro lado, T. glaucocolpa não foi incluída em nenhum dos trabalhos que utilizou dados moleculares, enquanto que a posição de T. cyanoptera continua ainda não é clara. O complexo de espécies T. episcopusT. sayacaT. glaucocolpa inclui 18 subespécies e uma grande variação morfológica, além de uma ampla distribuição de várias delas, com áreas de sintopia entre T. episcopus e T. sayaca, onde a identificação destas duas espécies é muito difícil. Estudos moleculares prévios só incluíram amostras de dois indivíduos de T. episcopus e uma de T. sayaca. Este complexo de espécies ainda apresenta uma grande instabilidade taxonômica. Na revisão deste gênero foram analisados 1.171 espécimes. As análises morfométricas mostraram que a massa é o parâmetro que apresenta a maior variação e que T. cyanoptera é a única mais claramente diferençável dentre as unidades taxonômicas. Foi também realizada uma análise filogenética com base em dois marcadores mitocondriais (Cyt- e ND2), além de três íntrons nucleares (íntron 3 do gen MUSK, íntron 5 do gen TGFB2 e uma parte do íntron 5 do gen BF5). Foi realizada uma análise RAxML e das redes de haplótipos independentemente para cada lócus, e esta informação foi utilizada para agrupar as amostras em unidades taxonômicas genéticas. O RAxML e as redes de haplótipos mostraram uma relação próxima entre T. episcopus e T. sayaca, além de uma alta probabilidade de um processo de introgressão entre as espécies. Há também uma evidente estruturação genética em T. episcopus. As unidades taxonômicas genéticas foram utilizadas em uma análise multilocus de árvore de espécies, com um relógio molecular calibrado. A árvore de espécies sugere que a origem do gênero Thraupis se deu entre 5.5 e 7.7 milhões de anos. Thraupis glaucocolpa é a linhagem mais antiga do gênero e a estrutura genética dentro de T. episcopus possui uma relação com as características morfológicas dessa espécie. Finalmente, são sinonimizadas várias subespécies e T. episcopus cana é elevada à espécie com base nos dados morfológicos e moleculares.

<p>Management of wolves is a complex issue, and molecular genetics is an important tool in this work. Molecular genetics can provide important information at the species, population and individual level, which can be essential for the development of management programs aiming at the long term survival of wolf populations.</p><p>In this thesis I developed new genetic markers on the canine Y chromosome to estimate the number of founders of the Scandinavian wolf population. This knowledge is important to reconstruct the history of the population and to design the most appropriate conservation strategies. Next, genetic markers with different pattern of inheritance have been used to identify hybrids between wolves and dogs. This allowed us to determine the direction of hybridization and to evaluate its possible impact on the gene pool of a wolf population. Furthermore, I also developed a method for a more reliable identification of the predator responsible of an attack by using saliva remains left on the prey. Since predation on livestock is perhaps the main reason for the negative opinions about the predator, the correct identification of the responsible for an attack (wolf, dog or hybrid) is essential. </p><p>Finally, this thesis has also been focusing on the domestication of dogs. By using Y chromosome markers (paternally inherited), it has been possible to complement previous studies based on mtDNA sequences (maternally inherited) and autosomal markers (inherited from both parents). In this way I have obtained a more complete picture of the domestication process and of the origin of breeds. This has shown that there has been a bias in the contribution of the two sexes in the origin of dog breeds (fewer males then females contributing to each breed) and that the origin of dogs was not marked by extensive backcrosses with male wolves over the entire species range.</p>

Management of wolves is a complex issue, and molecular genetics is an important tool in this work. Molecular genetics can provide important information at the species, population and individual level, which can be essential for the development of management programs aiming at the long term survival of wolf populations. In this thesis I developed new genetic markers on the canine Y chromosome to estimate the number of founders of the Scandinavian wolf population. This knowledge is important to reconstruct the history of the population and to design the most appropriate conservation strategies. Next, genetic markers with different pattern of inheritance have been used to identify hybrids between wolves and dogs. This allowed us to determine the direction of hybridization and to evaluate its possible impact on the gene pool of a wolf population. Furthermore, I also developed a method for a more reliable identification of the predator responsible of an attack by using saliva remains left on the prey. Since predation on livestock is perhaps the main reason for the negative opinions about the predator, the correct identification of the responsible for an attack (wolf, dog or hybrid) is essential. Finally, this thesis has also been focusing on the domestication of dogs. By using Y chromosome markers (paternally inherited), it has been possible to complement previous studies based on mtDNA sequences (maternally inherited) and autosomal markers (inherited from both parents). In this way I have obtained a more complete picture of the domestication process and of the origin of breeds. This has shown that there has been a bias in the contribution of the two sexes in the origin of dog breeds (fewer males then females contributing to each breed) and that the origin of dogs was not marked by extensive backcrosses with male wolves over the entire species range.

Although polyploidy is believed to be a major source of evolutionary novelty, few studies have examined the genetic basis of phenotypic variation in wild polyploids. In this thesis I have studied the genetic basis of flowering time variation in the wild tetraploid crucifer Capsella bursa-pastoris, as well as the evolutionary history of this species. First, phylogenetic methods were employed to test hypotheses on the origin of C. bursa-pastoris. Based on DNA sequences from two chloroplast DNA loci and three independent nuclear genes, we found no support for the notion of C. bursa-pastoris as an autopolyploid of the diploid C. grandiflora, or an allopolyploid of C. grandiflora and C. rubella, even though some C. bursa-pastoris accessions shared alleles with C. rubella at nuclear loci. Using divergence population genetic methods, a larger sample of accessions and data for six duplicated nuclear genes, we found that allele sharing in sympatry was better explained by introgressive hybridization than by multiple origins of the tetraploid. The genetic basis of flowering time variation was examined using three approaches. A gene expression microarray study revealed that early- and late-flowering accessions differ in circadian rhythm, as well as in the gibberellin pathway affecting flowering time. Second, two QTL (Quantitative Trait Loci) for flowering time map to duplicated linkage groups. Third, polymorphisms at the candidate genes CRYPTOCHROME1 (CRY1), in one of the QTL regions, and FLOWERING LOCUS C (FLC) are associated with natural flowering time variation. Different FLC splice site polymorphisms are associated with flowering time in samples from Western Eurasia and China. The CRY1 association is only found in Europe, where alleles introgressed from C. rubella have an effect on flowering time. In conclusion, duplicated genes, introgressive hybridization and splicing variation may all have played a role in the evolution of flowering time variation in C. bursa-pastoris.

Mémoire numérisé par la Division de la gestion de documents et des archives de l'Université de Montréal