Dissertations / Theses on the topic 'Microsatelliti nucleari'

The common octopus, Octopus vulgaris Cuvier, 1797, represents an important commercial fishery marine resource all over the world, characterized by increasing requests from the markets. In the Mediterranean Sea, especially for the Italian fishery, this species constitutes an important portion of trawling and artisanal landings. In Italy, despite the great socio-economic interest, specific management measures do not exist at the national level for this resource. The common octopus is a very interesting topic both for applied studies and for the basic research, especially at the taxonomic level, because of the uncertainties regarding the occurrence of a single species, with cosmopolitan distribution, or the existence of multiple species forming a ‘species complex’. The principal aim of this work is the genetic characterization of seven populations of O. vulgaris along the coasts of Sardinia in order to obtain useful indications for the selection of correct management units. The genetic analyses have been realized using three different genetic markers, two mitochondrial (the Cytochrome Oxidase I (COI) and III (COIII) genes), and one nuclear marker (five microsatellites loci). All markers proved to be useful in investigating the genetic structure of the populations. In particular, the mitochondrial sequences were useful in comparisons with homologous ones from the GenBank database to evaluate the species’ taxonomy. The results of AMOVA show a substantial homogeneity in Sardinian populations that display low levels of differentiation, both with the nuclear marker (FST=0.004 ns), and the mitochondrial ones (ΦSTCOI=0.003 ns; ΦSTCOIII=0.0002 ns). The pairwise analyses show low levels of differentiation and not significant values for all the genetic markers. The lack of significant genetic differentiation in the Sardinian samples is further confirmed by DAPC, PCA analyses and the Bayesian clustering of the software STRUCTURE. The demography was investigated using all the genetic markers, which pointed out the lack of demographic changes in recent time. The multimodal mismatch distributions reaffirm the occurrence of populations demographically stable. The stability of the populations was confirmed by the haplotype network analyses with the two mitochondrial markers, highlighting a common situation in stationary populations: several principal haplotypes, shared by all the locations, and an increasing number of new secondary haplotypes arising from mutational events. The COI and COIII sequences permitted the comparison of the Sardinian haplotypes with the O. vulgaris sequences available in GenBank. Both markers highlight a genetic affinity among Sardinian specimens and the sequences from the Mediterranean Sea (France, Spain, Central Mediterranean and Turkey), the Eastern Atlantic Ocean (Morocco, Senegal and Galicia), the Southern Atlantic Ocean (South Africa and Tristan da Cunha) and the Southern Indian Ocean (Amsterdam and Saint Paul Islands). This substantial genetic homogeneity contrasts with some sequences (from specimens collected in Turkey, Japan, China, Brazil and Venezuela) that resulted to be highly divergent from all the others. This finding reaffirms the potential existence of several O. vulgaris populations, just partially interconnected, or the even occurrence of distinct species, and emphasizes the need for more detailed phylogeographic and taxonomic studies of the Octopus genus to confirm or exclude the presence of cryptic species within the taxon, as suggested by several authors.

Sequences of the cytochrome b gene and genotypes from 11 polymorphic microsatellite loci were used to assess phylogeographic variation in ferruginous pygmy-owls (Glaucidium brasilianum) from Arizona, Mexico, and Texas. Analysis of mtDNA indicated that pygmy-owl populations in Arizona and Texas are unique, with no shared haplotypes. Populations from Sonora and Sinaloa, Mexico, were distinct from remaining populations in Mexico and grouped closest to haplotypes in Arizona. Nested clade analysis of mtDNA sequence data indicated past fragmentation separated pygmy-owls into two major groups: 1) Arizona, Sonora and Sinaloa, Mexico, and 2) southwestern (Nayarit and Michoacan), south-central (Oaxaca and Chiapas), and eastern Mexico, along the eastern slope of the Sierra Madre Oriental from Texas to Central America. In addition, analysis of mtDNA variation in several species of Glaucidium support the recommendation that populations of G. brasilianum from Mexico, Texas, and Arizona represent a phylogenetically distinct group from populations occurring in South America. The level of separation between the North and South Americanpopulations justifies granting species status (G. ridgwayi) to the North American population. Analysis of distance matrices derived from genotypes of 11 polymorphic microsatellite loci supports restricted gene flow between pygmy-owl populations in Arizona-Sonora and Sinaloa, and Texas-Tamaulipas and the remainder of states in Mexico. The Arizona-Sonora population showed signs of a recent genetic bottleneck, an observation supported by low population estimates for Arizona (13-117 individuals). Heterozygosity in Arizona, however, was equal to levels recorded throughout Mexico and Texas. Congruent patterns revealed by both mtDNA and nuclear DNA (microsatellites) indicate Arizona and Texas populations are distinct subspecies that require the design and implementation of separate management plans for recovery and conservation efforts.

Microsatellites are repetitive DNA characterized by tandem repeats of short motifs (2 – 5 bp). High mutation rates make them ideal for population level studies. Microsatellite allele genesis is generally attributed to strand slippage, and it is assumed that alleles are caused only by changes in repeat number. Most analyses are limited to alleles (electromorphs) scored by mobility only, and models of evolution rarely account for homoplasy in allele length. Additionally, insertion/deletion events (indels) in the flanking region or interruptions in the repeat can obfuscate the accuracy of genotyping. Many investigators use microsatellites, designed for a focal species, to screen for genetic variation in non-focal species. Comparative studies have shown different mutation rates of microsatellites in different species, and even individuals. Recent studies have used reciprocal comparisons to assess the level of polymorphism of microsatellites between pairs of taxa. In this study, I investigated the evolution of microsatellites within a phylogenetic context, using comparisons within the rodent family Bathyergidae. Bathyergidae represents a monophyletic group endemic to sub-Saharan Africa and relationships are well supported by morphological and molecular data. Using mitochondrial and nuclear DNA, a robust phylogeny was generated for the Bathyergidae. From my results, I proposed the new genus, Coetomys. I designed species-specific genotyping and microsatellite flanking sequence (MFS) primers for each genus. Sequencing of the MFS provided direct evidence of the evolutionary dynamics of the repeat motifs and their flanking sequence, including rampant electromorphic homoplasy, null alleles, and indels. This adds to the growing body of evidence regarding problems with genotype scores from fragment analysis. A number of the loci isolated were linked with repetitive elements (LTRs and SINEs), characterized as robust phylogenetic characters. Results suggest that cryptic variation in microsatellite loci are not trivial and should be assessed in all studies. The phylogenetic utility of the nucleotide variation of the MFS was compared to the well-resolved relationships of this family based on the 12S/TTR phylogeny. Variation observed in MFS generated robust phylogenies, congruent with results from 12S/TTR. Finally, a number of the indels within the MFS provided a suite of suitable phylogenetic characters.

This study reexamines the nuclear microsatellite analysis by Rooney et al. (1999a) of Bering-Chukchi-Beaufort Seas bowhead whales (Balaena mysticetus) to determine if this population underwent a genetic bottleneck as a result of 19th and early 20th Century commercial whaling. This investigation used more accurate laboratory techniques to score alleles, had a larger sample size that was divided into two groups (mainland Alaska and St. Lawrence Island (SLI)), and used a moderately different set of microsatellite loci which are more variable and thus, more informative. The results corroborate the findings of Rooney et al. (1999a) for mainland Alaska showing no evidence of a genetic bottleneck. However, the SLI data analyses provide conflicting conclusions. The Wilcoxon test is significant for a heterozygote excess (p = 0.042) suggesting that a genetic bottleneck has occurred. This is not substantiated by the exact tests of each locus or the table-wide sign test. There is a possibility that a bottleneck has occurred, but due to the small sample size this is not a definitive conclusion and warrants reanalysis with a larger sample size.

Norway spruce is one of two dominating species in Swedish forestry and the most economically important tree species in Sweden. In order to preserve the ability to adapt to a changing environment and to keep populations healthy, genetic diversity has to be preserved. When choosing a small number of individuals from a natural stand to establish a seed orchard the population size decrease. With only a small number of genetically different individuals the risk of inbreeding increase. Furthermore if many clones of the same tree are used in one seed orchard there is also an increased risk selfing. The aim of this study was therefore to investigate whether genetic diversity in Norway spruce differs between age groups and if this can be attributed to forestry practices. All sampling was done from a single location in Västerbotten, Sweden and the different age groups were chosen to represent stands not affected by the modern forest industry to recently planted forests. The chosen age groups are young (12-18 years), intermediate (30-45 years), and old (above 85 years). From each age group 150 individuals were sampled. With genomic microsatellite markers each individual was genotyped at eight simple sequence repeat (SSR) loci. Results show an overall high genetic diversity with an average expected heterozygosity (He) at 0.842 and low genetic differentiation with an average fixation index among populations (FST) of 0.003. The genetic diversity of each age group was also high (He 0.832 to 0.843) and the inbreeding coefficient ranged from 0.061 in the old group to 0.078 in the intermediate group. The pairwise FST value was highest between the old group and the young group but the differentiation was only 0.005 (P=0.001). An analysis of molecular variance also showed that only 0.34% of the total genetic variance was explained by differences among age groups. This study found little evidence for a decrease in genetic diversity due to forestry practices and revealed high genetic diversity and low differentiation between the age groups, indicating a healthy population.

The structuring of Atlantic salmon (Salmo salar L.) into discrete, genetically differentiated populations both within and between river catchments is well documented. The utilisation of this knowledge has proved valuable in a variety of evolutionary, ecological, managerial and conservation contexts. In this thesis, the genetic structuring of Atlantic salmon populations in northwest Europe was assessed in two catchments of very different sizes, using a range of molecular and associated population genetic methods; findings from the catchment level research are set in context by a broader phylogeographic study of post-glacial colonisation of the region. A regional study into the glacial origins and post-glacial colonisation routes of Atlantic salmon in northwest Europe was explored by analysing a pre-existing microsatellite dataset and supplementing it with haplotype data from mtDNA PCR-RFLP analysis of the same samples (N=702). Evidence from allele permutation tests undertaken on the microsatellite data alongside mtDNA haplotype frequencies suggested that there was a cryptic northern refuge in northwest France, with colonisation of the British Isles and Ireland occurring from this and the long-known Iberian Peninsula refuge. Catchment level studies were undertaken on the river Dart and river Tweed, involving 1151 fish being genotyped with 14 microsatellite loci with a subset of 211 fish being genotyped by mtDNA PCR-RFLP. In both catchments, populations were found to be weakly differentiated genetically, and were most consistent with the meta-population theory of evolution. Similarly, individual spatial autocorrelation analysis indicated that each major tributary within the catchments could be considered as a distinct management or conservation unit. In the Tweed dataset, however, limitations in the sample coverage across the catchment reduced the robustness of some findings. Historical stocking of the river Dart with fish from Scotland and Iceland is well-documented. The long-term implications of these activities on contemporary Dart populations were assessed by genotyping 177 fish from the donor populations using scale samples taken in the 1960s and comparing them to contemporary Dart populations by undertaking admixture analysis. Overall, admixture between the donor and recipient populations was low and appeared to reflect natural underlying levels of genetic relationships. However, increased admixture of donor stocks with one extant Dart population was apparent, indicating some potentially long-term localised success of the stocked fish through hybridisation with the native populations; nevertheless, with the population continuing to decline, this should not be viewed as a successful supplementation programme. Two tributaries on the river Tweed, the Gala and Leader, were inaccessible to salmon for long periods due to the construction of barriers to migration. On both tributaries, fish passes were installed in the 1940s and re-colonisation of the tributaries was possible. Assignment analysis was undertaken and indicated that, contrary to findings for between catchment studies, salmon straying from the most proximate tributaries (i.e. the Ettrick and Caddon) did not appear to be the principal colonisers of the current Gala and Leader populations. Rather, the highest proportion of Gala samples assigned to the Teviot (42%), with the Leader populations assigning to many tributaries across the catchment (Ettrick 28%; Upper 21%; Teviot 19%). However, given the relatively weak differentiation of the baseline samples and limitations inherent in the dataset, the correct self-assignment of baseline samples was very low (average 26%; range 0-47%), hence interpretation must be undertaken with caution. Nevertheless, the findings suggest that the Gala population may have reached a temporally stable state in the 60 years since it has been accessible to salmon. Whilst the relatively small scale of these studies is acknowledged, the application of the findings in management and conservation of the species are discussed in a wider context. These studies would support the following recommendations: to include information on the historic (refugial) origin of contemporary populations in regional management strategies; to treat each major tributary as a distinct unit as an appropriate scale for catchment level management; and, with stocking and supplementation programmes appearing to have no significant long-term success, coupled with the relative speed with which extirpated tributaries appear to be naturally re-colonised, the use of stocking and supplementation programmes should be discouraged.

O Brasil é o país mais rico do mundo em espécies de psitacídeos (cerca de 74), sendo 17 delas ameaçadas de extinção. Entre elas está a arara-azul (Anodorhynchus hyacinthinus) que é considerada vulnerável e pode se tornar ameaçada num futuro próximo, em conseqüência do intenso tráfico ilegal e perda do seu habitat. No presente estudo estimamos os níveis de variabilidade e caracterizamos a estrutura genética de populações naturais de A. hyacinthinus. Analisamos 10 locos de microssatélites de 98 indivíduos e seqüências concatenadas de genes mitocondriais (ND5, citocromo-b e ND2; 2123 pb total) de 80 indivíduos. O índice de diversidade genética foi considerado baixo em relação a outras espécies de psitacídeos. Além disso, os índices RST e a análise bayesiana dos dados de microssatélites indicaram moderada estruturação genética entre indivíduos de quatro regiões geográficas (Pantanal norte, Pantanal sul, norte e nordeste), mas os índices de FST indicaram diferenciação somente entre três regiões (norte e nordeste sem diferenciação). A estruturação entre essas três regiões foi congruente com a forte estruturação genética apontada pelos índices de FST e pela rede de haplótipos das seqüências mitocondriais. Baseado nos dados mitocondriais o tempo de divergência entre os grupos genéticos de A. hyacinthinus foi estimado em 16 a 42 mil anos atrás, o que corresponde ao final do Pleistoceno. Ainda, os resultados apontaram para uma população demograficamente estável ao longo do tempo, o que pode indicar que a baixa variabilidade pode ser uma característica da espécie. Entretanto, a rede de haplótipos apresenta forma em estrela com alguns haplótipos de baixa freqüência, o que pode indicar expansão recente, principalmente para região nordeste. Baseado nos dados de estruturação genética populacional, foi possivel indicar a possível origem de indivíduos apreendidos e sem procedência conhecida, o que é importante para realizar ações preventivas de repressão e fiscalização. Adicionalmente, foram analisados sete locos de microssatélites de filhotes amostrados no mesmo ninho (mesma estação reprodutiva, estações reprodutivas consecutivas e estações alternadas) em duas regiões do Pantanal. Os resultados sugerem que a espécie é predominantemente monogâmica estrita, mas há pelo menos 12,5% de paternidade extra-par e 6,5% de parasitismo de ninho. Além disso, foram confirmados dados obtidos em campo de que muitos casais utilizam o mesmo ninho em anos consecutivos e alternados. Finalmente, padronizamos a sexagem molecular de amostras de penas de muda. Concluindo, os resultados genéticos obtidos nesse trabalho trazem informações sobre os processos envolvidos na história evolutiva dessa espécie, além de contribuir com informações sobre o comportamento reprodutivo das araras-azuis proporcionando mais subsídios para elaboração de programas de conservação.
Brazil has the highest number of parrot species in the world (about 74), 17 of them endangered. Among them is the hyacinth macaw (Anodorhynchus hyacinthinus), which is considered vulnerable and could become endangered in the near future, due to the intense illegal traffic and loss of habitat. In this study we estimated levels of variability and characterized the genetic structure of natural populations of hyacinth macaws. We analyzed 10 microsatellite loci from 98 individuals and concatenated sequences of mitochondrial genes (ND5, cytochrome b and ND2, 2,123 bp total) from 80 individuals. The genetic diversity index was low compared to those from other species of parrots. In addition, RST indeces and Bayesian analysis of microsatellite data showed moderate genetic structure among individuals of four regions in Brazil (north Pantanal, south Pantanal, north and northeast), but FST indeces indicate differentiation only between three regions (north and northeast without differentiation). This is in accordance with the strong genetic structure indicated by FST indeces and haplotype network based on mitochondrial sequences. Based on the mitochondrial data, the time of divergence of the genetic groups of hyacinth macaws was estimated to have occurred 16 to 42 thousand years ago, which corresponds to the late Pleistocene. Still, the results suggest that the population has been demographically stable over time, which may indicate that the low variability levels may be a characteristic of the species. However, the haplotype network presents a star shape, which indicate recent expansion, specially in the northeast. Additionally, given the population genetic structure data, it was possible to identify the most probable region of origin of apprehended individuals, this information is important to plan preventive and repressive control. Additionally, we analyzed seven microsatellite loci of chicks sampled in the same nest (same breeding season, alternate breeding seasons and consecutive seasons) in two regions of the Pantanal. The results suggest that the species is predominantly monogamous, but there is at least 12.5% of extra-pair paternity and 6.5% of brood parasitism. Furthermore, the genetic data is congruent with field observations that suggest that many couples return to the same nest in consecutive and alternative breeding seasons. Finally, we standardized for a molecular sexing protocol for molten feathers. In conclusion, the genetic results obtained in this study provide information about the processes involved in the evolutionary history and the reproductive behavior of hyacinth macaws that may help plan conservation actions.

The feral pig, Sus scrofa, is a widespread and abundant invasive species in Australia. Feral pigs pose a significant threat to the environment, agricultural industry, and human health, and in far north Queensland they endanger World Heritage values of the Wet Tropics. Historical records document the first introduction of domestic pigs into Australia via European settlers in 1788 and subsequent introductions from Asia from 1827 onwards. Since this time, domestic pigs have been accidentally and deliberately released into the wild and significant feral pig populations have become established, resulting in the declaration of this species as a class 2 pest in Queensland. The overall objective of this study was to assess the population genetic structure of feral pigs in far north Queensland, in particular to enable delineation of demographically independent management units. The identification of ecologically meaningful management units using molecular techniques can assist in targeting feral pig control to bring about effective long-term management. Molecular genetic analysis was undertaken on 434 feral pigs from 35 localities between Tully and Innisfail. Seven polymorphic and unlinked microsatellite loci were screened and fixation indices (FST and analogues) and Bayesian clustering methods were used to identify population structure and management units in the study area. Sequencing of the hyper-variable mitochondrial control region (D-loop) of 35 feral pigs was also examined to identify pig ancestry. Three management units were identified in the study at a scale of 25 to 35 km. Even with the strong pattern of genetic structure identified in the study area, some evidence of long distance dispersal and/or translocation was found as a small number of individuals exhibited ancestry from a management unit outside of which they were sampled. Overall, gene flow in the study area was found to be influenced by environmental features such as topography and land use, but no distinct or obvious natural or anthropogenic geographic barriers were identified. Furthermore, strong evidence was found for non-random mating between pigs of European and Asian breeds indicating that feral pig ancestry influences their population genetic structure. Phylogenetic analysis revealed two distinct mitochondrial DNA clades, representing Asian domestic pig breeds and European breeds. A significant finding was that pigs of Asian origin living in Innisfail and south Tully were not mating randomly with European breed pigs populating the nearby Mission Beach area. Feral pig control should be implemented in each of the management units identified in this study. The control should be coordinated across properties within each management unit to prevent re-colonisation from adjacent localities. The adjacent rainforest and National Park Estates, as well as the rainforest-crop boundary should be included in a simultaneous control operation for greater success.

En plus des facteurs environnementaux et démographiques, les propriétés génétiques des populations sont devenues une préoccupation majeure pour préserver les populations en déclin de l'extinction. Afin d’acquérir des informations pertinentes pour la planification et la mise en œuvre des stratégies de conservation, les biologistes de la conservation ont réalisé le besoin d’avoir des connaissances en génétique des populations. Grace à l'acquisition de plus en plus rapide et de moins en moins chère d'une large gamme de marqueurs moléculaires, le recours a l’usage de l’outil moléculaire se répand de plus en plus. Ainsi, la génétique de la conservation se confirme comme une discipline à part entière qui est donc l’utilisation de la génétique dans la préservation des espèces comme entités dynamiques capables d'évoluer pour faire face aux changements environnementaux et afin de minimiser leur risque d'extinction. Par le biais de l’utilisation d’un large panel de marqueurs moléculaires (gènes mitochondriaux et nucléaires, microsatellites et SNPs), nous nous sommes intéresse à l’histoire évolutive à différentes échelles spatio-temporelles de la sous-espèce ouest méditerranéenne Testudo. hermanni hermanni (THH), qui présente une distribution insulaire et continentale très fragmentée. Le but de ce travail consiste à 1) comprendre les processus qui expliqueraient la distribution actuelle de la diversité génétique des populations et leur structure, 2) identifier l'origine des populations introduites (à Minorque et au Delta de l’Ebre), et 3) dater l’origine de la sous espèce THH. A l’échelle des populations, il s’agit d’identifier le nombre de groupes génétiques homogènes chez la tortue d’Hermann et le degré de différentiation génétique entre ces groupes afin de définir des unités de conservation évolutivement significatives (ESU) et des unités de gestion (MU). Enfin, nous nous sommes intéresses à l’étude des derniers noyaux de populations de THH dans le Var par des approches de génétique du paysage. Nos résultats ont révélé qu’une divergence par vicariance est à l’ origine de l’apparition de la sous-espèce T.h. hermanni. Ce scenario biogéographique s’expliquerait par les successions d’évènements glaciaires et interglaciaires qu’a connu le Pléistocène depuis plus de 2 MA provoquant un mouvement de retrait de l’espèce vers des zones refuges sur la frange côtière nord-méditerranéenne. Par ailleurs, le patron de différentiation mitochondriale Ile-continent observe et confirme par les microsatellites est très original par rapport à ce qui est connu chez d’autres espèces de reptiles partageant la même aire de distribution. Au vue de l’analyse phylogénétique confirmée par les microsatellites, on peut affirmer que la tortue d’Hermann n’est pas native sur Minorque et qu’elle a une double origine : la première, résultant d’une introduction à partir d’une seule source, probablement d’une population continentale génétiquement proche des Albères. La seconde d'origine insulaire, serait le résultat d’apports multiples, à partir de la Corse, de la Sardaigne ou de la Sicile. Enfin, l’isolement des populations de THH au sein de chaque région géographique reflète une structure génétique très forte. Par conséquent, six unités de gestion (MUs) sont proposées comme unités de conservation et de suivi sur le terrain
In addition to environmental and demographic factors, the study of genetic properties of populations became inevitable issues in the conservation of declining populations. To acquire relevant information for conservation planning and implementing conservation strategies, conservationists have realized the need of population genetics tools. Moreover, this discipline has become more efficient with the development of a wide range of effective and relatively cheap methods for the characterization of a huge number of molecular markers. This led to define the conservation genetics as a separate discipline, which is the use of genetics in species preservation as dynamic entities evolving to cope with environmental changes and to minimize their extinction risk. Using a broad panel of molecular markers (mitochondrial and nuclear genes, microsatellites and SNPs), we interested in the evolutionary history at different spatial and temporal scales of the Mediterranean western subspecies Testudo hermanni hermanni (THH), which presents a very fragmented insular and continental distribution. The aim of this study is to 1) understand the processes that explain the current distribution of the structure and genetic diversity of populations, 2) identify the origin of introduced populations (Menorca and Ebro Delta) and 3) Dating the origin of the subspecies THH. At the population level, our study aimed to identify the number of homogeneous genetic groups of THH tortoise and the degree of genetic differentiation between these groups in order to identify evolutionarily significant units (ESU) and management units (MU). Finally, we were interested in the study of the last core populations of THH in the Var by landscape genetics approach. Our results revealed that a divergence by vicariance pattern explains the origin of the appearance of the subspecies THH. This biogeographic scenario is explained by the succession of glacial and interglacial events of the Pleistocene causing a withdrawal of the species toward refugia on the northern Mediterranean fringe. Moreover, the observed differentiation pattern (island vs continent) is very original compared to the reported diversity patterns of other reptiles sharing the same distribution range. According to our results, we may conclude that the Hermann’s tortoise is not native in Menorca and has a double origin: the first, is an introduction resulting from a unique source, probably from a continental lineage genetically close to Albera. The second, from an island origin, is the result of multiple contributions, from Corsica, Sardinia or Sicily. Lastly,the isolation of THH populations within each geographic region reflects a very strong genetic structure, therefor the six most relevant management units forconservation purposes are proposed on the basis that they represent a significant part of the evolutionary legacy of the species

Effects of man made pollutants on an ecosystem are initiated at the cellular level where a prime determinant for survival of an organism is its ability to metabolise and excrete toxic chemicals or their metabolites, thereby preventing cellular toxicity or damage to germ cell DNA. Cytochrome P450 (CYP) enzymes are responsible (in concert with the remainder of the Ah battery enzymes) for the metabolism of numerous xenobiotics and endogenous compounds, including the metabolic activation of most environmental toxic chemicals and carcinogens. Genetic polymorphisms which affect performance of these enzymatic detoxification systems may alter tolerance to pollutants and thus survival in polluted environments. Alterations in the susceptibility of individuals and the development of resistant populations has arisen by forced selection of populations with variant genes, resulting in increased detoxification capacity. There is evidence for such scenarios of variations in activities of pollutant biotransforming enzymes of fish contributing to survival in polluted estuarine environments and several chemically resistant populations have been identified in the USA and Europe. In fish it has been demonstrated that CYP1A enzyme activity is required to activate some carcinogenic xenobiotics to a metabolic state in which they can form DNA adducts. The mechanism of reduced CYP1A expression in highly contaminated populations may therefore represent resistance to chemical stressors. European flounder (Platichthys flesus) from some waterways which have a long history of severe sedimentary contamination do not show elevated levels of CYP1A. The aim of the current study was to investigate whether any heritable differences were apparent between offspring from parents inhabiting long-term polluted and pristine areas. Flounder were obtained from a highly polluted estuary in the UK and crossed with fish from a relatively pristine environment. Offspring were raised in communal tanks in order to standardise environmental conditions, and allow investigations into the genetic variation of CYP1A. To allow identification of offspring to parental fish, polymorphic microsatellite loci were isolated and characterised for the flounder. Novel cDNA probes to transcription factors in the detoxification pathway (AhR2 and ARNT2) were cloned for flounder, and RT-PCR / Southern blot methods were developed for quantitation of gene transcript levels. A novel method of CYP1A quantification using real-time PCR was developed. PAH and PCB exposure trials were carried out on mixed batch offspring, and CYP1A gene transcript levels assessed using Northern blot and real-time PCR techniques. Offspring were genotyped to their parents using the microsatellites obtained, and CYP1A transcript levels were correlated with clean and polluted areas. CYP1A was further correlated to transcription factor expression, and data are presented. Following exposure to the commercial PCB mixture, Aroclor 1254, CYP1A transcript levels were found to be significantly lower in families whose parents originated from a polluted area. This observation indicates that there is a possible genetic component to variation in CYP1A levels, and that these fish may have acquired a heritable tolerance to polluted areas. The lack of induction, or correlation with CYP1A levels, of AhR2 and ARNT2 expression indicates a possible AhR independent pathway for the metabolism of PCBs in the flounder. © Tom Dixon 2003 http://www.tomdixon.org

Wu, Feng.
Thesis (M.Phil.)--Chinese University of Hong Kong, 2010.
Includes bibliographical references (leaves 38-44).
Abstracts in English and Chinese.
Title Page --- p.i
Thesis Committee --- p.ii
Acknowledgment --- p.iv
Table of Contents --- p.v
List of Figures --- p.vii
List of Abbreviations and Symbols --- p.xi
Abstract (English version) --- p.xii
Abstract (Chinese version) --- p.xiii
Chapter 1 --- Introduction --- p.1
Chapter 1.1 --- Significance of DNA CCTG repeats --- p.1
Chapter 1.2 --- Objectives of this work --- p.2
Chapter 1.3 --- DNA structure --- p.3
Chapter 2 --- Materials and Methods --- p.5
Chapter 2.1 --- Sample design --- p.5
Chapter 2.2 --- Sample preparation --- p.5
Chapter 2.3 --- NMR spectroscopy --- p.6
Chapter 2.4 --- Resonance assignment --- p.7
Chapter 3 --- NMR Structural Studies of (CCTG)3 --- p.9
Chapter 3.1 --- Overview --- p.9
Chapter 3.2 --- NMR resonance assignments --- p.9
Chapter 3.3 --- Formation of two-residue CT-loop in the middle repeat of (CCTG)3 --- p.12
Chapter 3.4 --- C-bulge and T.T mispair in (CCTG)3 hairpin stem region --- p.13
Chapter 3.5 --- Summary --- p.15
Chapter 4 --- NMR Structural Studies of (CCTG)4 --- p.16
Chapter 4.1 --- Overview --- p.16
Chapter 4.2 --- Conformational exchange in (CCTG)4 --- p.16
Chapter 4.3 --- Formation of two-residue CT-loops in different repeats of (CCTG)4 --- p.17
Chapter 4.4 --- Mutational studies of (CCTG)4 --- p.19
Chapter 4.4.1 --- Mutational studies on the 1st repeat of (CCTG)4: (CCTG)4-C2T --- p.19
Chapter 4.4.2 --- Mutational studies on the 2nd repeat of (CCTG)4：(CCTG)4-C6T --- p.21
Chapter 4.4.3 --- Mutational studies on the 3rd repeat of (CCTG)4：(CCTG)4-C 10T --- p.26
Chapter 4.4.4 --- Mutational studies on the 4th repeat of (CCTG)4: (CCTG)4-C14T --- p.28
Chapter 4.5 --- Summary --- p.33
Chapter 5 --- Conclusions and Future Works --- p.35
References --- p.38

Red pine ( Pinus resinosa Ait.) is an ecologically and economically important forest tree species of northeastern North America and is considered as one of the most genetically depauperate conifer species in the region. Traditional genetic markers have failed to identify significant genetic polymorphism in this species. In order to set conservation priorities and gain insight into the post-glacial dispersal history of red pine, highly sensitive markers such as nuclear microsatellites are valuable. I have isolated and characterized thirteen nuclear microsatellite loci by screening a partial genomic library with di-, tri-, and tetra-nucleotide repeat oligonucleotide probes. Analysis of 518 individuals representing 17 red pine populations from Manitoba through Newfoundland identified five polymorphic microsatellite loci with an average of 9 alleles per locus. The mean expected and observed heterozygosity values were 0.508 and 0.185 respectively. Private alleles were detected in six of the populations examined, and F coefficients showed significant departure from Hardy-Weinberg equilibrium with an excess of homozygosity indicating high levels of inbreeding in all populations studied. Populations were highly differentiated with approximately 31.5% of genetic variation among populations, and results assuming IAM and SMM were similar. Weak but significant isolation by distance was detected, and genetic distances revealed regional genetic breaks and distinct populations of red pine. These findings suggest a complex and highly differentiated population genetic structure for red pine uncharacteristic of most conifers. The genetic patterns identified support a multiple refugia hypothesis for red pine, which should be taken into consideration when setting conservation plans for this species.

碩士
國立臺灣大學
生態學與演化生物學研究所
100
Fagus hayatae Palib., a rare and statutory-conserved plant in Taiwan, is distributed in Chatienshan Nature Reserve, Tongshan area, and few new single mountain areas. It can also be found in Sichun province, Hubei province, and Zhejiang province in China. A population genetics research on this endangered species was conducted by using chloroplast trnL–F and rbcL–atpB regions; three microsatellite markers developed for Fagus crenata were also examined in Fagus hayatae on the nine populations in Taiwan and three populations in China. Results showed that a low variation in Taiwan''s populations using chloroplast markers, and a close relationship with the related taxa from China. Results also indicated a clear divergence of the populations from Syue Mountain (Beicha, Loupei, Lala, Niauzui, and Ayu) and Central Mountains (Lenkan, Dabai, Taipin Beech Trail, and Tongsan). By conducting microsatellite analysis, populations in Taiwan exhibited low level of genetic variation, while populations in China had higher level of genetic variation, especially for Zhejian’s population. Nevertheless, results of Bayesian clustering analysis suggested that populations in Taiwan and China should be considered as a single genetically distinct group. The results from microsatellite data provide more genetic information and therefore can be a great resource for further population genetics studies, hopefully provide applications to conservation biology of Fagus hayatae in Taiwan.

The number of non-parasitic (brook) lamprey species in the genus Lampetra is underestimated since isolated populations are generally considered one species due to their relatively conserved body form. The phylogeographic and phylogenetic structure was estimated among and within Lampetra species along the Pacific coast of North America (presumed to represent Lampetra richardsoni; L. pacifica – which is currently regarded as a junior synonym of L. richardsoni; L. ayresii; and L. hubbsi) using up to three mitochondrial and three nuclear genetic markers. These data show that L. richardsoni as currently recognized is polyphyletic when lampreys (some of which show up to 8 K2P% sequence divergence) from Siuslaw River and Fourmile Creek (Oregon) and Mark West, Paynes, and Kelsey creeks (California) are included; Lampetra pacifica is a valid species; the population from Kelsey Creek almost certainly represents a new species; and those from Siuslaw, Fourmile, and Mark West may also be distinct species.