Asexual modes of reproduction are usually based on the principle of copying (cloning) DNA from the female and passing it on to the offspring. For most asexually reproducing vertebrates the progeny develop from an unreduced and often unfertilised egg. This is driven by the mechanisms of parthenogenetic and gynogenetic reproduction. While in the former the clonal germ cell develops spontaneously and separately, in the latter a sexual partner is needed to activate the cleavage of the ovum, although without the fusion of the sperm and egg. Therefore in both cases there is no fertilization and the clonal progeny consist solely of daughters, hence the majority of previous studies have only focused on asexual female lineages. However, on rare occasions asexual clonal males can arise when the right fertilization occurs. Whilst these offspring are usually infertile, fertile diploid asexual males have been discovered in just three genera of hybrid origin in vertebrates. One of these unique cases is the European water frog complex of the genus Pelophylax, whose distribution includes the Czech Republic. In areas around the upper Odra River populations of hybrid males were recently discovered who form stable all-male lineages, similar to those formed by asexual females. The results of this study show that males produce...

The aim of this work was the construction of set of microsatellites for the Grey Partridge (Perdix perdix). I succeeded in isolation of seven new specific microsatellites. Another microsatellite markers were passed from close relative galliform species (Galliformes). On the whole I gained 18 microsatellite loci which were used for study of population charakteristics of three partridge populations from Stodůlky, Písecko and Milešín. The analysis showed conspicuous structuredness of these populations and surprisingly low values (less then 100) of their effective population sizes. Seven of the microsatellites were chosen for analysis of paternity. Indeed I found the evidence of extrapair paternity in this putatively monogamous bird. Finally I conclude, on the basis of sequenation of mitochondrial control region that our populations belong to the western linage of mitochondrial DNA.

No description available.

Genealogy reconstruction is widely used in biology when relationships among entities are studied. Phylogenies, or evolutionary trees, show the differences between species. They are of profound importance because they help to obtain better understandings of evolutionary processes. Pedigrees, or family trees, on the other hand visualize the relatedness between individuals in a population. The reconstruction of pedigrees and the inference of parentage in general is now a cornerstone in molecular ecology. Applications include the direct infer- ence of gene flow, estimation of the effective population size and parameters describing the population’s mating behaviour such as rates of inbreeding. In the first part of this thesis, we construct genealogies of various types of cancer. Histopatho- logical classification of human tumors relies in part on the degree of differentiation of the tumor sample. To date, there is no objective systematic method to categorize tumor subtypes by maturation. We introduce a novel algorithm to rank tumor subtypes according to the dis- similarity of their gene expression from that of stem cells and fully differentiated tissue, and thereby construct a phylogenetic tree of cancer. We validate our methodology with expression data of leukemia and liposarcoma subtypes and then apply it to a broader group of sarcomas and of breast cancer subtypes. This ranking of tumor subtypes resulting from the application of our methodology allows the identification of genes correlated with differentiation and may help to identify novel therapeutic targets. Our algorithm represents the first phylogeny-based tool to analyze the differentiation status of human tumors. In contrast to asexually reproducing cancer cell populations, pedigrees of sexually reproduc- ing populations cannot be represented by phylogenetic trees. Pedigrees are directed acyclic graphs (DAGs) and therefore resemble more phylogenetic networks where reticulate events are indicated by vertices with two incoming arcs. We present a software package for pedigree reconstruction in natural populations using co-dominant genomic markers such as microsatel- lites and single nucleotide polymorphism (SNPs) in the second part of the thesis. If available, the algorithm makes use of prior information such as known relationships (sub-pedigrees) or the age and sex of individuals. Statistical confidence is estimated by Markov chain Monte Carlo (MCMC) sampling. The accuracy of the algorithm is demonstrated for simulated data as well as an empirical data set with known pedigree. The parentage inference is robust even in the presence of genotyping errors. We further demonstrate the accuracy of the algorithm on simulated clonal populations. We show that the joint estimation of parameters of inter- est such as the rate of self-fertilization or clonality is possible with high accuracy even with marker panels of moderate power. Classical methods can only assign a very limited number of statistically significant parentages in this case and would therefore fail. The method is implemented in a fast and easy to use open source software that scales to large datasets with many thousand individuals.:Abstract v Acknowledgments vii 1 Introduction 1 2 Cancer Phylogenies 7 2.1 Introduction..................................... 7 2.2 Background..................................... 9 2.2.1 PhylogeneticTrees............................. 9 2.2.2 Microarrays................................. 10 2.3 Methods....................................... 11 2.3.1 Datasetcompilation ............................ 11 2.3.2 Statistical Methods and Analysis..................... 13 2.3.3 Comparison of our methodology to other methods . . . . . . . . . . . 15 2.4 Results........................................ 16 2.4.1 Phylogenetic tree reconstruction method. . . . . . . . . . . . . . . . . 16 2.4.2 Comparison of tree reconstruction methods to other algorithms . . . . 28 2.4.3 Systematic analysis of methods and parameters . . . . . . . . . . . . . 30 2.5 Discussion...................................... 32 3 Wild Pedigrees 35 3.1 Introduction..................................... 35 3.2 The molecular ecologist’s tools of the trade ................... 36 3.2.1 3.2.2 3.2.3 3.2.1 Sibship inference and parental reconstruction . . . . . . . . . . . . . . 37 3.2.2 Parentage and paternity inference .................... 39 3.2.3 Multigenerational pedigree reconstruction . . . . . . . . . . . . . . . . 40 3.3 Background..................................... 40 3.3.1 Pedigrees .................................. 40 3.3.2 Genotypes.................................. 41 3.3.3 Mendelian segregation probability .................... 41 3.3.4 LOD Scores................................. 43 3.3.5 Genotyping Errors ............................. 43 3.3.6 IBD coefficients............................... 45 3.3.7 Bayesian MCMC.............................. 46 3.4 Methods....................................... 47 3.4.1 Likelihood Model.............................. 47 3.4.2 Efficient Likelihood Calculation...................... 49 3.4.3 Maximum Likelihood Pedigree ...................... 51 3.4.4 Full siblings................................. 52 3.4.5 Algorithm.................................. 53 3.4.6 Missing Values ............................... 56 3.4.7 Allelefrequencies.............................. 58 3.4.8 Rates of Self-fertilization.......................... 60 3.4.9 Rates of Clonality ............................. 60 3.5 Results........................................ 61 3.5.1 Real Microsatellite Data.......................... 61 3.5.2 Simulated Human Population....................... 62 3.5.3 SimulatedClonalPlantPopulation.................... 64 3.6 Discussion...................................... 71 4 Conclusions 77 A FRANz 79 A.1 Availability ..................................... 79 A.2 Input files...................................... 79 A.2.1 Maininputfile ............................... 79 A.2.2 Knownrelationships ............................ 80 A.2.3 Allele frequencies.............................. 81 A.2.4 Sampling locations............................. 82 A.3 Output files..................................... 83 A.4 Web 2.0 Interface.................................. 86 List of Figures 87 List of Tables 88 List Abbreviations 90 Bibliography 92 Curriculum Vitae I

No description available.

Examination of variation at polymorphic microsatellite loci is a widely accepted method for determining parentage and examining genetic diversity within rainbow trout (Oncorhynchus mykiss) breeding programs. Genotyping costs are considerable; therefore, we developed a single-step method of co-amplifying twelve microsatellite loci in two hexaplex reactions. The protocol is explicitly described to ensure reproducible results. I applied the protocol to samples previously analyzed at the National Center for Cool and Coldwater Aquaculture (NCCCWA) with previously reported marker sets for a comparison of results. Each marker within the multiplex system was evaluated for duplication, null alleles, physical linkage, and probability of genotyping errors. Data from four of the 12 markers were excluded from parental analysis based on these criteria. Parental assignments were compared to those of a previous study that used five independently amplified microsatellites. Percentages of progeny assigned to parents were higher using the subset of eight markers from the multiplex system than with five markers used in the previous study (98% vs. 92%). Through multiplexing, use of additional markers improved parental allocation while also improving efficiency by reducing the number of PCR reactions and genotyping runs required. I evaluated the methods further through estimation of F-statistics, pairwise genetic distances, and cluster analysis among brood-years at the NCCCWA facility. These estimates were compared to those from nine independently amplified microsatellites used in a previous study. Fst metrics calculated between brood-years showed similar values of genetic differentiation using both marker sets. Estimates of individual pairwise genetic distances were used for constructing neighbor-joining trees. Both marker-sets yielded trees that showed similar subpopulation structuring and agreed with results from a model-based cluster analysis and available pedigree information. These approaches for detecting population substructure and admixture portions within individuals are particularly useful for new breeding programs where the founders' relatedness is unknown. The 2005 NCCCWA brood-year (75 full-sib families) was challenged with Flavobacterium psychrophilum, the causative agent of bacterial coldwater disease (BCWD). The overall mortality rate was 70%, with large variation among families. Resistance to the disease was assessed by monitoring post-challenge days-to-death. Phenotypic variation and additive genetic variation were estimated using mixed models of survival analysis. The microsatellite markers used were previously isolated from BAC clones that harbor genes of interest and mapped onto the rainbow trout genetic linkage map. A general relationship between UBA gene sequence types and MH-IA-linked microsatellite alleles indicated that microsatellites mapped near or within specific major histocompatibility (MH) loci reliably mark sequence variation at MH genes. The parents and grandparents of the 2005 brood-year families were genotyped with markers linked to the four MH genomic regions (MH-IA, MH-IB, TAP1, and MH-II) to assess linkage disequilibrium (LD) between those genomic regions and resistance to BCWD. Family analysis suggested that MH-IB and MH-II markers are linked to BCWD survivability. Tests for disease association at the population level substantiated the involvement of MH-IB with disease resistance. The impact of MH sequence variation on selective breeding for disease resistance is discussed in the context of aquaculture production. / Master of Science

No description available.

L’huître creuse, Crassostrea gigas, est une espèce dont la production aquacole représente un intérêt économique tant au niveau mondial qu’aux niveaux européen et français. Cependant, cette espèce subit des mortalités estivales enregistrées dès le début du 20ème siècle et, depuis 2008, ce phénomène s’est amplifié et menace essentiellement les huîtres juvéniles. La production aquacole d’huître creuse subit les conséquences de ces mortalités massives ; c’est pourquoi ce phénomène est étudié depuis de nombreuses années. En France, la bactérie Vibrio splendidus et le virus Ostreid Herpes Virus 1 (OsHV-1) sont le plus souvent associés aux épisodes de mortalités massives d’huîtres creuses juvéniles et il a été démontré que les individus sélectionnés pour leur résistance aux mortalités estivales étaient capables de ralentir l’augmentation de la charge virale en OsHV-1 dans leurs tissus puis de la faire régresser. Ces mêmes individus présenteraient également un effort reproducteur plus modeste que des individus sélectionnés pour leur sensibilité aux mortalités estivales. Ce travail de thèse s’inscrit dans ce contexte et a donc eu pour principal objectif d’améliorer la connaissance de l’architecture génétique de la reproduction de C. gigas en identifiant des régions du génome ou QTLs (Quantitative Trait Loci) impliquées dans l’effort reproducteur et de mettre en évidence d’éventuelles relations génétiques entre survie et reproduction, des QTLs impliqués dans la survie ayant déjà été détectés. Afin de caractériser l’effort reproducteur, il a été nécessaire de développer un ensemble de nouveaux outils. D’un point de vue biologique, 21 familles F2 ont été produites à partir des lignées sélectionnées pour leur réponse contrastée aux mortalités estivales. D’un point de vue moléculaire, de nouveaux marqueurs SNP (Single Nucleotide Polymorphism) ont été développés afin d’augmenter la densité de la carte génétique déjà disponible pour C. gigas. D’un point de vue technique, l’Imagerie par Résonance Magnétique (IRM) a permis d’observer la gamétogenèse de 300 individus d’une même famille F2 au cours de huit sessions réparties sur deux années alors que les études précédentes étaient limitées à une observation ponctuelle; les méthodes classiques d’observation de la gamétogenèse entrainant nécessairement la mort des animaux. Une forte corrélation a été mise en évidence entre les observations par IRM et par la méthode classique de l’histologie. En plus de l’estimation du rapport gonado-somatique (indice traditionnellement utilisé pour caractériser l’effort reproducteur), l’IRM a également permis d’observer des variations individuelles de la cinétique de la gamétogenèse ainsi que des différences entre les mâles et les femelles; le sexe étant identifiable sur les images obtenues par IRM. Parallèlement, 300 individus de deux autres familles F2 ont été sacrifiés pour estimer le rapport gonado-somatique par histologie. Cette approche a ainsi permis de détecter des QTLs impliqués dans de nombreux traits concernant la gamétogenèse. Des individus provenant des trois familles F2 caractérisés pour l’effort reproducteur ont été caractérisés pour la survie à un épisode de mortalités estivales. Cette étude a permis de détecter des QTLs impliqués dans le caractère « survie ». Ces QTLs correspondent, pour certains, à ceux détectés au cours d’une étude précédente. De plus, ces QTLs sont parfois colocalisés avec des QTLs impliqués dans l’effort reproducteur. Bien que la reproduction de l’huître creuse soit un caractère complexe à suivre, les nouveaux outils utilisés au cours de ce travail de thèse ont permis d’acquérir de nouvelles connaissances. Le séquençage du génome complet de Crassostrea gigas ainsi que les nouvelles méthodes de séquençage pourront peut-être permettre d’affiner les régions QTLs détectées. / The Pacific oyster, Crassostrea gigas, is a major aquacultured species whose production represents an economic interest at worldwide, european and french levels. However, this species undergoes summer mortalities recorded from the beginning of the 20th century and, since 2008, this phenomenon increased and threatens mainly juvenile oysters. Aquaculture production of oysters suffers consequences of mass mortalities, that’s why this phenomenon has been studied for many years. In France, the bacterium Vibrio splendidus and the Ostreid virus Herpes Virus 1 (OsHV-1) are often associated with mass mortality outbreaks of juveniles oysters and it was demonstrated that selected individuals for resistance to summer mortality were able to slow the increasing in viral load OsHV-1 in their tissues and then to decline it. These same individuals also present a lighter reproductive effort than individuals selected for their sensitivity to summer mortality. In this context, this study aimed to improve the knowledge of genetic architecture of reproduction of C. gigas by identifying some regions of the genome called QTLs (Quantitative Trait Loci) involved in reproductive effort and highlighting possible genetic relationships between reproduction and survival; QTLs involved in survival being already detected. To characterize the reproductive effort, it was necessary to develop a set of new tools. From a biological point of view, 21 F2 families were produced from lines selected for their contrasting response to summer mortality. From a molecular point of view, new SNPs (Single NucleotidePolymorphism) were developed to increase density of the genetic map already available for C. gigas. On a technical point of view, Magnetic Resonance Imaging (MRI) allowed to observe the gametogenesis of 300 individuals of the same family F2 during eight sessions over two years while previous studies were limited to a one-time observation because of the conventional methods of observation of gametogenesis leading necessarily to the death of the animals. A strong correlation was found between observations by MRI and observations by the conventional method of histology. In addition to the estimation of gonadic index (index traditionally used to characterize there productive effort), MRI also revealed individual variations in kinetics of gametogenesis and differences between males and females, the sex being identifiable on MRI images. In parallel, 300 individuals from two F2 families were sacrificed to estimate the gonadic index by histology. This approach enabled the detection of QTLs involved in many gametogenesis traits. Individuals from the three families characterized for F2 reproductive effort were characterized for survival during a summer mortality outbreak. This study was able to detect QTLs involved in the trait "survival". These QTLs correspond to some of those detected in a previous study. In addition, these QTLs are often collocated with QTLs involved in reproductive effort. Although there production of the Pacific oyster is a complex trait to follow, the new tools used in this thesis allowed acquiring new knowledges. The sequencing of genome of Crassostrea gigas and Next-Generation Sequencing technologies may be able to help to refine the detected QTL regions.

La perchaude (Perca flavescens) constitue une ressource socioéconomique de grande importance dans le Lac Saint-Pierre (Québec, Canada). Bien que ce lac fluvial soit désigné réserve de la biosphère par l’UNESCO, le statut de la perchaude est préoccupant. Afin de permettre à l’espèce de persister en fonction des diverses pressions anthropiques, il est important de comprendre sa dynamique populationnelle et les mécanismes qui en sont responsables. La perchaude est connue pour sa philopatrie ; le fait de toujours se reproduire à son site de naissance peut entraîner la subdivision d’une espèce en de multiples populations, où chacune pourra être pourvue d’adaptations locales distinctes. Il est possible d’étudier ces processus à l’aide des signaux génétiques associés à la reproduction des individus. Toutefois, une faible différentiation génétique entre les populations du Lac Saint-Pierre est envisagée en raison de la colonisation récente du système (moins de 8000 ans). L’objectif de cette étude est de déterminer s’il existe plusieurs populations de perchaude dans le Lac Saint-Pierre. Les simulations réalisées ont révélé que l’utilisation de marqueurs AFLP (amplified fragment length polymorphism), permettant une analyse globale du génome, affiche une meilleure détection de la différentiation des populations que celle des marqueurs microsatellites. Afin d’associer les individus à leur site de naissance, la méthode d’AFLP et des microsatellites ont été utilisées sur des larves capturées suite à l’éclosion des oeufs. Trois analyses distinctes d’AFLP ont indiqué une corrélation entre la composition génétique des individus et des sites géographiques, confirmant ainsi la présence de plusieurs populations sympatriques dans le Lac Saint-Pierre, découlant vraisemblablement de la philopatrie de l’espèce. L’absence de différentiation génétique relatée par les marqueurs microsatellites vient confirmer l’importance du choix des marqueurs génétiques. Bien que la différentiation génétique observée soit relativement faible, la gestion de la perchaude devrait tenir compte de la dynamique des populations distinctes dans ce système. / Yellow perch (Perca flavescens) is a commercially-exploited freshwater fish species in Lake Saint-Pierre (Quebec, Canada). Even if this fluvial lake is designated as a biosphere reserve by UNESCO, the yellow perch status is of concern. To ensure the persistence of species facing to anthropogenic pressures, understanding dynamics and mechanism that structure populations are of major importance. Because the habitat characteristics are spatially structured and this species is known to display natal site fidelity, a subdivision of the species in multiple populations and local adaptations may occur. It is possible to detect these processes according to genetic signal associated with individuals’ reproduction. However, low genetic differentiation is hypothesised due to the recent colonization of the system (< 8 K years). This study aims at determining if there are multiple populations of yellow perch in Lake Saint-Pierre. Simulations was first performed to confirm that population differentiation is better depicted by amplified fragment length polymorphism (AFLP) than microsatellite markers. In order to associate individuals to their site of birth, larvae captured after hatching were used. A survey of the variation throughout the entire genome was then performed by using the AFLP approach and variations at microsatellite loci were used to further investigate the organization of these populations. Three distinct AFLP analyses indicated a correlation between genetic composition of individuals and spawning sites, and thus confirmed the presence of multiple sympatric populations in Lake Saint-Pierre, resulting likely from natal site fidelity. At the opposite, the lack of genetic differentiation reported by microsatellites markers confirms the importance of the choice of genetic markers. While the genetic differentiation is very low, the management of this species should take into account the existence of distinct population structures in this system.

A Mata Atlântica é considerada uma das grandes prioridades para a conservação de biodiversidade em todo o continente americano. Este bioma é um dos oito hotspots do planeta e caracteriza-se pelo alto grau de endemismo e pela alta diversidade de espécies que encerra. As abelhas Euglossini estão restritas à região Neotropical, com uma faixa de distribuição que se estende desde o Norte do México até a província de Córdoba, na Argentina. As espécies dessa tribo são abundantes em florestas tropicais úmidas/chuvosas e florestas subtropicais da América Central e América do Sul onde ocorrem em maior diversidade. Em virtude do alto impacto causado pelas ações humanas em áreas de Mata Atlântica e o pouco que se conhece sobre sua diversidade, este trabalho propõe um estudo geral sobre a estrutura das comunidades de espécies de Euglossini presentes em uma área de planície litorânea continental, Parque Estadual da Serra do Mar - Núcleo Picinguaba (PESM), e uma área insular, Parque Estadual da Ilha Anchieta (PEIA), no município de Ubatuba, norte de São Paulo, Brasil. O PESM abrange uma área de aproximadamente 47.500 hectares e quase todos os ecossitemas da Mata Atlântica são encontrados em seus domínios. O PEIA cobre toda a extensão da Ilha Anchieta (828 hectares), distante cerca de 600 m do continente. As coletas foram realizadas mensalmente, de Agosto/2007 a Julho/2009, com o uso de 14 compostos aromáticos utilizados como iscas artificiais para a atração de machos, além de indivíduos coletados em flores. Mil quinhentos e setenta e seis espécimes (73 fêmeas e 1503 machos), pertencentes a 24 espécies, foram registrados no presente trabalho. A espécie mais abundante foi Euglossa cordata (36,4%), seguida por Euglossa iopoecila (13,9%) e Euglossa sapphirina (13,3%). Dezoito espécies foram amostradas na Ilha Anchieta, sendo que quatro delas foram exclusivas desta área, ao passo que 20 espécies foram amostradas no PESM e seis delas foram registradas apenas neste local. Na Ilha Anchieta, Eg. cordata representou 63,2%, quase dois terços do total de espécies coletadas. Já em Picinguaba, Eg. iopoecila (23,0%) e Eg. sapphirina (21,0%) foram as duas espécies dominantes. A espécie Eg. iopoecila, a mais abundante na área continental, não foi registrada na llha e Eg. sapphirina, a segunda espécie mais frequente no PESM, foi representada apenas por nove indivíduos na Ilha Anchieta e por 200 em Picinguaba. Estes dados sugerem que essas duas espécies podem atuar como bioindicadores de ambientes preservados, como sugerido para outras espécies de Euglossini. Por outro lado, alguns autores evidenciaram que Eg. cordata é uma espécie favorecida por ambientes perturbados, o que pode estar associado à sua alta abundância na Ilha Anchieta. Dados de literatura mostram que há um sinal evidente de perturbação ambiental, além da perda de espécies, é a dominância de algumas espécies. A Ilha Anchieta apresenta um longo histórico de ocupação humana e diversas espécies de plantas e animais foram introduzidos em seus domínios. / The Atlantic Forest is considered as one of the highest priorities for the conservation of biodiversity in the American continent. This biome is one of the eight biodiversity hotspots in the world and is characterized by high species diversity and endemism. Euglossini bees are restricted to the Neotropics, with a geographic range that extends from northern Mexico to the Córdoba Province in Argentine. Species from this tribe are abundant in humid and rainy tropical forests and in sub-tropical forests of Central and South America, where their diversity is high. Due to the high impact caused by human activities in the Atlantic Forest and the poor knowledge on its biodiversity, in the present study we assessed the community structure of the Euglossini in a coastal lowland area: Parque Estadual da Serra do Mar - Núcleo Picinguaba (PESM); and in an insular area: Parque Estadual da Ilha Anchieta (PEIA), Ubatuba, both located in northern São Paulo State, Brazil. PESM has an area of approximately 47.500 ha and almost all ecosystems of the Atlantic Forest are found within its domains. PEIA covers all the extension of the Anchieta Island (828 ha), about 600 m away from the continent. Sampling was carried out monthly, from August 2007 to July 2009, using artificial baits with 14 aromatic compounds to attract males, as well as by collecting individuals on flowers. One thousand five hundred and seventy-six specimens (73 females and 1,503 males), represented by 24 species were recorded. The most abundant species was Euglossa cordata (36.4%), followed by Euglossa iopoecila (13.9%) and Euglossa sapphirina (13.3%). Eighteen species were sampled on Anchieta Island and four of them were exclusive to this area; whereas 20 species were sampled in PESM and six of them were only recorded there. On Anchieta Island, E. cordata represented almost two thirds of the total species collected (63.2%). In Picinguaba, E. iopoecila (23.0%) and E. sapphirina (21.0%) were the dominant species. E. iopoecila, the most abundant species in the continental area, was not recorded on the island, and E. sapphirina, the second most frequent species in PESM, was represented only by nine individuals on Anchieta Island and by 200 in Picinguaba. Results suggest that these two species may act as bioindicators of preserved environments, as suggested for other Euglossini species. However, some authors evidenced that E. cordata is favored by disturbed environments, which may be associated with its high abundance on Anchieta Island. Other studies show that an evident sign of environmental perturbation, besides species loss, is the dominance of some species. Anchieta Island exhibits a long history of human occupation and several plant and animal species were introduced there.