Breeding programmes that use elite sires with the best estimated breeding values for muscling traits have achieved significant improvement in lamb production in the UK. Further acceleration of the rate of genetic gain for the desirable production traits could be achieved using DNA marker-assisted selection (MAS) breeding strategies. The underlying causal genetic variants associated with improved muscling may be unknown and lying between a cluster of genes known as quantitative trait loci (QTL) or could be single nucleotide polymorphisms (SNP). LoinMAXTM, Texel muscling QTL (TM-QTL) and c.*1232G > A myostatin mutation were genetic variants that reported to be associated with improved muscling characteristics and hence subjected to further analysis in this project. It is essential before incorporating segregating genetic variants in any breeding scheme to comprehensively evaluate their effects on carcass traits. In-vivo scanning (ultrasound scanning (US) and computed tomography scanning (CT)), and carcass video image analyses (VIA) were used in the current studies. Objective VIAprediction weights of the carcass primal cuts could be the backbone of a value-based marketing system that is suggested to replace the current Meat and Livestock Commission (MLC) carcass grades for conformation scores (MLC-C) and fat class (MLC-F). The effect of a single copy of LoinMAXTM QTL (LM-QTL) compared to noncarriers was evaluated in UK crossbred lambs out of Scottish Mule ewes. M. longissimus lumborum (MLL) width, depth and area, as measured by CT scanning, were significantly greater in lambs heterozygous for LM-QTL compared to noncarriers. VIA detected a significant effect of the LM-QTL on the predicted weight of saleable meat yield in the loin primal cut (+2.2%; P < 0.05). The effects of the ovine c.*1232G > A myostatin mutation (MM), found on sheep chromosome 2, on carcass traits in heterozygous crossbred lambs sired by Texel and Poll Dorset rams were studied. Texel crossbred lambs carrying MM had increased loin depth and area. In both crossbred lambs, MM-carriers had significantly higher CT-estimated lean weight and proportion (2 to 4%) and muscle to bone ratios (by ~3%). Poll Dorset heterozygous crossbred animals had higher muscle to fat ratio (28%) and significantly lower fat-related measurements. The c.*1232G > A (MM) mutation as well as TM-QTL effects were evaluated in a different genetic background of Texel x Welsh Mountain crossbreed lambs. Carrying two copies of MM was associated with a significant positive effect on 8 week weight, a negative effect on ultrasound fat depth, a substantial decrease in MLC-fat score, positive impact on VIA-estimated weight of the hind leg, chump and loin primal cuts, as well as the muscularity of the hind leg and loin regions with greater loin muscle width, depth and area. Two copies of MM altered lambs‟ morphological traits with significantly wider carcasses across the shoulders, breast and hind legs and greater areas of the back view of the carcass when measured by VIA. TM-QTL significantly increased US-muscle depth and TM-QTL carriers had significantly greater loin muscle width and area measurements. Comparing TM-QTL genetic groups (homozygote allele carriers (TM/TM), heterozygote carriers of paternal and maternal origin of allele (TM/+ and +/TM, respectively) and homozygote non-carriers (+/+)) and TM-QTL mode of action were then studied. TM/TM carcasses were significantly heavier than non-carriers by 1.6 kg and scored higher conformation values when compared to heterozygote groups only. TM/+ lambs had significantly higher VIA-predicted weight and muscularity in the hind leg and loin, and higher loin dimensions relative to some other genotypic groups. The effect of TM-QTL on some carcass shape measurements was significant. TM-QTL mode of action results on the loin muscling traits supports the earlier reports of polar over dominance. In the light of growing calls to replace the current subjective carcass payment system with the objective VIA system that values the carcass according to the superiority of its cuts, I investigated the ability of US and CT measurements to predict the VIAestimated weights of the carcass primal cuts. Several prediction equations were examined but the best could be achieved when ultrasound measurement, CT linear measurements and live weight were fitted in the model. Since CT scanning information of elite sires is now being used for genetic selection for carcass merit, genetic parameters and genetic relationships between CT scanning measurements and post mortem traits (VIA and MLC-FC) were estimated. However, results were not sufficiently accurate to be of practical use due to lack of data.

Chicken is a major protein source and intensively selected for economically important traits by humans. As such, this generated a huge range of phenotypes that representing a diverse spectrum of genetic variation. Understanding the functional basis of the genetic variants that underlie these traits, however, remains a formidable endeavour particularly for complex traits. Nonetheless, molecular phenotyping of an organism from sequenced data is doable with the advances in bioinformatics analysis and unparalleled surveys of genome wide genetic variants. This provides the opportunity to gain insights into the genome architecture and assists in identifying chromosomal regions underlying selection through a “sequences to consequences” approach. Combining a whole genome re-sequencing (WGS) approach with the knowledge of selection history, this thesis aimed to study the chromosomal regions and genetic variants underlying traits of interest in various selected chicken populations. To achieve this, genetic (quantitative and population genetics), genomic and bioinformatics approaches were employed and integrated to investigate the genome wide selection signatures in a number of different lines of chicken selected for different complex traits. This includes analysing: (i) divergently selected broilers for fatness traits (Chapter 2), (ii) a closed population of layer chickens (Chapter 3), (iii) selection signatures unique to broiler or layer chickens (Chapter 4) and (iv) selection signatures in colony stimulating factor 1 (CSF1) associated with gene expression differences in broiler and layer populations (Chapter 5). Candidate genes and nucleotides underlying potential selection regions were identified, and attempts were made to further elucidate the potential interplay between genes and the biological pathways involved in regulating traits in these selected chicken lines. Incorporating integrative approaches, variants within selection signatures were annotated to provide further evidence of their functional consequences. Overall, non-coding regions were enriched in selection signatures implied that causative variants may have regulatory roles. Capitalising on the millions of genetic variants discovered from WGS, chromosomal regions subject to selection were detected using a number of population genetics statistics. In broiler chicken lines divergently selected for very low-density plasma lipoprotein (VLDL) (Chapter 2), incorporating signatures of selection helped to improve the resolution of previously mapped quantitative traits loci (QTL) intervals. This research demonstrated that the integration of the analysis of selection signatures with functional annotation of genetic variants enabled refinement and characterisation of the QTL for fatness traits. In a closed population of brown leghorn layers (Chapter 3), evidence of selection signatures was found through Tajima’s D analysis. The analysis unravelled selection signatures encoding genes involved in numerous pathways and genes having key roles such as in behaviour, including feather pecking. Combining population differentiation statistic (FST) and Tajima’s D, a number of regions subject to divergent selection between broilers and white egg layers were identified (Chapter 4). Selection signatures were found to harbour mutations involved in cellular and tissue development, including genes having important roles in growth, fatness, egg shell strength and muscle development. These regions and the overlapping genes thereby may be potentially contributing to the different phenotypic variations observed between broilers and layers. In Chapter 5, a revised gene model for colony stimulating factor 1 (CSF1) showing complex pattern of alternate transcripts was predicted from transcriptome analysis of RNA isolated from 21 different tissues. In parallel, selection signatures analysis with the FST statistic, identified selection signatures that differentiate broilers to white egg layers (3 regions) or brown egg layers to white egg layers (4 regions). All these selection signatures were located within non-coding regions, indicating potential divergent selection of CSF1 within regulatory regions. Overall, the results presented in this thesis using the “sequences to consequences” approach, link several genomic regions and genes to phenotypic variation in domesticated chicken lines. The work reported here serves as a foundation for further study to decipher the relationship between “genotype and phenotype” and its functional consequences due to selection.

Orientador: Lucia Galvão de Albuquerque / Resumo: Registros de características quantitativas e informações genotípicas cole- tadas para cada animal são utilizados para identificar regiões do genoma associadas à variação fenotípica. No entanto, essas investigações são, geralmente, realizadas com base em testes estatísticos de correlação ou associação, que não implicam em causalidade. A fim de explorar amplamente essas informações, métodos poderosos de inferência causal foram desenvolvidos para estimar os efeitos causais entre as variáveis estudadas. Apesar do progresso significativo neste campo, inferir os efeitos causais entre variáveis aleatórias contínuas ainda é um desafio e poucos estudos têm explorado as relações causais em genética quantitativa e no melhoramento animal. Neste contexto, dois estudos foram realizados com os seguintes objetivos: 1) Buscar as relações causais entre as características de carcaça e qualidade de carne usando um modelo de equação estrutural (MEE), sob modelo linear misto em bovinos da raça Nelore, e 2) Reconstruir redes de genes-fenótipos e realizar análise de rede causal por meio da integração de dados fenotípicos, genotípicos e transcriptômicos em bovinos da raça Nelore. Para o primeiro estudo, um total de 4.479 animais com informação fenotípica para o peso da carcaça quente (PCQ), área de olho lombo (AOL), espessura de gordura subcutânea (EGS), força de cisalhamento (FC) e marmoreio (MAR) foram usados. Os animais foram genotipados usando os painéis BovineHD Bead- Chip e GeneSeek Genomic Pro... (Resumo completo, clicar acesso eletrônico abaixo) / Abstract: Quantitative traits and genotypes information have been collected for each animal and used to identify genome regions related to phenotypes variation. However, these investigations are, usually, performed based on correlation or association statistical tests, which do not imply in causation. In order to fully explore these information, powerful causal inference methods have been developed to estimate causal effects among the variables under study. Despite significant progress in this field infer causal effect among random variables remains a challenge and some few studies have explored causal relationships in quantitative genetics and animal breeding. In this context, two studies were performed with the following objectives: 1) Search for the causal relationship among carcass yield and meat quality traits using a structural equation model (SEM), under linear mixed model context in Nelore cattle, and 2) Reconstruct gene-phenotype networks and perform causal network analysis through the integrating of phenotypic, genotypic, and transcriptomic data in Nelore cattle. For the first study, a total of 4,479 animals with phenotypic information for hot carcass weight (HCW), longissimus muscle area (LMA), backfat thickness (BF), Warner-Bratzler shear force (WBSF), and marbling score (MB) traits were used. Animals were genotyped using BovineHD BeadChip and GeneSeek Genomic Profiler Indicus HD - GGP75Ki. For causal inference using SEM a multistep procedure methodology was used as follow:... (Complete abstract click electronic access below) / Doutor

Von Willebrand Disease (VWD) is the most common bleeding disorder. In addition to known major genes, genetic modifiers, such as ABO blood group, affect quantitative outcome measures for VWD. The data consist of an 854-member Amish pedigree with established linkage of VWD to a locus within the Von Willebrand Factor (VWF) gene on chromosome 12. The DNA sequence of the causative mutation is known. Phenotypic information and genotypic data consisting of VWF mutation status and a genome screen of markers are available for 385 pedigree members. Genetic modifiers of the VWF mutation are investigated using known and new conditional linkage methods that search for modifier genes of a major gene with known mutation. The MCMC-based program LOKI was used to conduct multipoint linkage analysis of VWD outcome measures while controlling for the VWF mutation. Adjustment for the mutation did not eliminate the linkage signal on chromosome 12 in the same location as the VWF mutation. Evidence for QTLs was also found on six other chromosomes. Smod, a score statistic that detects evidence of a genetic modifier conditional on linkage to a major gene, was developed for sib pair data. To limit the modifier gene main effect, Smod was developed so that variance due to the modifier locus is bounded above by the variance of the interaction between major gene and modifier gene. The performance of Smod was compared to other published score statistics. Power to detect linkage to the modifier locus depended on major gene and modifier gene risk allele frequencies, relative contribution of the major gene main effect to the interaction effect, and the upper bound on the modifier gene main effect. The Amish pedigree was broken up into sib pair data and analyzed using Smod and other score statistics. Using these statistics, the strongest evidence for QTLs for VWD was also found on chromosome 12 in the region of the VWF mutation. Combined with the LOKI results, further analysis will help determine if intragenic modification is occurring or if linkage disequilibrium between the mutation and analyzed markers is driving results.

This project investigated the genetic composition of natural populations of Anadara trapezia in Australia at three spatial scales : i) microgeographic (within an estuary, 50 metres to ~ 6 kilometres); ii) microgeographic (within populations, less than 50 metres); and, iii) macrogeographic (hundreds of kilometres along the coast of Australia). Allozyme polymorphism surveys using cellulose acetate strips have revealed, from 43 enzymes screened, 18 putative polymorphic loci. Comparisons of levels of heterozygosity among enzyme structural groups showed no significant differences, however, monomers were significantly more variable as a group than multimers. Significant differences in the level and distribution of polymorphism among functional groups of enzymes were observed. It appears that selection may be acting at the molecular level, not only on a particular locus, but on a group of functionally similar loci. At the macrogeographic scale, significant departures from random mating were observed in most populations. Significant differences in allele distribution among populations of A. trapezia along the east coast of Australia were found. At the macrogeographic scale, heterogeneity of allele frequencies may depend upon the distance separating the populations and surface water currents. Differentiation among population groups in this study is attributed to changes in the direction of the East Australian Current combined with onshore countercurrents. The systematic status of the disjunct western Australian population of A. trapezia was also evaluated as compared with the east coast populations. No evidence of genetic, hence evolutionary divergence was found. The results have serious implications in the management of fisheries as erroneous assumptions in fisheries management models may lead to depletion and near extinction of marine species. The research stresses the necessity of sampling at multiple scales and replication strategies. It also highlighted the complexities researchers are faced with in studies of marine bivalves, such as the presence of null alleles, deficiencies of heterozygotes, apparent inbreeding and the small geographic scales governing population structure. / Doctor of Philosophy (PhD)

Paget's disease of the bone is a skeletal disorder of unknown cause. This disease is characterised by excessive and abnormal bone remodelling brought about by increased bone resorption followed by disorganised bone formation. Increased bone turnover results in a disorganised mosaic of woven and lamellar bone at affected skeletal sites. This produces bone that is expanded in size, less compact, more vascular, and more susceptible to deformity or fracture than normal bone. Symptoms of Paget's disease may include bone pain, bone deformity, excessive warmth over bone from hypervascularity, secondary arthritis, and a variety of neurologic complications caused in most instances by compression of the neural tissues adjacent to pagetic bone. Genetic factors play a role in the pathogenesis of Paget's disease but the molecular basis remains largely unknown. The identification of the molecular basis of Paget's disease is fundamental for an understanding of the cause of the disease, for identifying subjects at risk at a preclinical stage, and for the development of more effective preventive and therapeutic strategies for the management of the condition. With this in mind, the aim of this project is to identify genetic loci, in a large pedigree, that may harbour genes responsible for Paget's disease of bone. A large Australian family with evidence of Paget's disease was recruited for these studies (Chapter 3). This pedigree has characterised over 250 individuals, with 49 informative individuals affected with Paget's disease of bone, 31 of whom are available for genotypic analysis. The pattern of disease in these individuals is polystotic, with sites of involvement including the spine, pelvis, skull and femur. Although the affected individuals have a severe early-onset form of the disease, the clinical features of the pedigree suggest that the affected family members have Paget's disease and not familial expansile osteolysis (a disease with some similarities to Paget's disease), as our patients have extensive skull and axial skeletal involvement. The disease is inherited as an autosomal dominant trait in the pedigree with high penetrance by the sixth decade. Due to the large size of this family and multiple affected members, this pedigree is a unique resource for the detection of the susceptibility gene in Paget's disease. The first susceptibility loci for Paget's disease of bone have been mapped by other investigators to chromosome 6p21 (PDB1) and 18q21.1-q22 (PDB2) in different pedigrees. Linkage analysis of the Australian pedigree in these studies was performed with markers at PDB1: these data showed significant exclusion of linkage, with LOD scores < - 2 in this region (Chapter 4). Linkage analysis of microsatellite markers from the PDB2 region excluded linkage with this region also, with a 30 cM exclusion region (LOD score < -2.0) centred on D18S42 (Chapter 4). This locus on chromosome 18q21.1-q22 contains a serine protease (serpin) cluster with similarities to chromosome 6p21. Linkage analysis of this region also failed to provide evidence of linkage to this locus (Chapter 4). These data are consistent with genetic heterogeneity of Paget's disease of bone. A gene essential for osteoclast formation encoding receptor activator of nuclear factor-kB (RANK), TNFRSF11A, has been previously mapped to the PDB2 region. Mutations in the TNFRSF11A gene have been identified segregating in pedigrees with Familial Expansile Osteolysis and early onset familial Paget's disease, however, linkage studies and mutation screening have excluded the involvement of RANK in the majority of Paget's disease patients. For the Australian pedigree, mutation screening at the TNFRSF11A locus revealed no mutations segregating with affected individuals with Paget's disease (Chapter 4). Based on these findings, our hypothesis is that a novel susceptibility gene relevant to the pathogenesis of Paget's disease of bone lies elsewhere in the genome in the affected members of this pedigree; this gene should be identifiable using a microsatellite genome-wide scan followed by positional cloning. A genome-wide scan of the Australian pedigree was carried out, followed by fine mapping and multipoint analysis in regions of interest (Chapter 5). The peak 2-point LOD scores from the genome-wide scan were LOD = 2.75 at D7S507 and LOD = 1.76 at D18S70. Two additional regions were also considered for fine mapping: chromosome 19p11-q13.1 with a LOD of 1.58 and chromosome 5q35-qter with a LOD of 1.57. Multipoint and haplotype analysis of markers flanking D7S507 did not support linkage to this region (Chapter 5). Similarly, fine mapping of chromosome 19p11-q13.1 failed to support linkage to this region (Chapter 5). Linkage analysis with additional markers in the region on chromosome 5q35-qter revealed a peak multipoint LOD score of 6.77 (Chapter 5). A distinct haplotype was shown to segregate with all members of the family, except the offspring of III-5 and III-6. Haplotype analysis of markers flanking D18S70 demonstrated a haplotype segregating with Paget's disease in a large sub-pedigree (descendants of III-3 and III-4) (Chapter 5). This sub-pedigree had a significantly lower age at diagnosis than the rest of the pedigree (51.2 + 8.5 vs. 64.2 + 9.7 years, p = 0.0012). Linkage analysis of this sub-pedigree demonstrated a peak two-point LOD score of 4.23 at marker D18S1390 (q = 0.00), and a peak multipoint LOD score of 4.71, at marker D18S70. An implication of these data is that 18q23 harbours a novel modifier gene for reducing the age of onset of Paget's disease of bone. A number of candidate Paget's genes have previously been identified on chromosome 18q23, including the nuclear factor of activated T cells (NFATc1), membrane-associated guanylated kinase (MAGUK) and a zinc finger protein. Candidate gene sequencing of these genes in these studies has failed to identify mutations segregating with affected family members in the sub-pedigree linked to chromosome 18q23 (Chapter 6). More recently, a mutation in the gene encoding the ubiquitin-binding protein sequestosome 1 (SQSTM/p62) has been shown to segregate with affected members of Paget's disease families of French-Canadian origin. In this study, a single base pair deletion (1215delC) was identified as segregating with the majority of affected members in the pedigree (Chapter 6). This deletion introduces a stop codon at amino acid position 392 which potentially results in early termination of the protein and loss of the ubiquitin binding domain. The three affected members of the family that do not share the affected haplotype do not carry a mutation in the coding region of SQSTM/p62. Screening of affected members from 10 further Paget's disease families identified the previously reported P392L mutation in 2 (20%) families. No SQSTM1/p62 coding mutations have been found in the remaining 8 families or in 113 aged matched controls. In conclusion, this project has identified genetic loci and mutations that segregate with individuals affected with Paget's disease. Further investigation of the functional significance of the genetic changes at these loci is expected to lead to a better understanding of the molecular basis of this disease.

Needle-to-stem unit rate (NESTUR) is a stem growth index of conifer seedling trees that measures the efficiency of stemwood production per unit of needle growth. Five experiments were carried out in this thesis using progenies of two unrelated full-sib radiata pine crosses. The initial experiment (experiment 1) applied the bulked segregant analysis technique to determine whether RAPD analysis could be successfully extended to the development of molecular markers for NESTUR in radiata pine. The NESTUR values of 174 progenies of the full-sib family 12038 x 10946 were determined. Based on the genotypic analysis of the individuals, two quantitative trait loci (QTL) controlling NESTUR were identified at ANOVA P-levels of 0.01-0.001. An absence of RAPD fragment markers generated by primers OPE-06 and OPA-10 was associated with low NESTUR values, while primer UBC-333 generated a 550 bp band that was associated with high NESTUR values. Linkage to components of NESTUR (increments in stem diameter and stem volume) was demonstrated for one of the QTL, while the other was unique to NESTUR, and not shared with the components. There was a significant interaction between the two QTLs. Presence of OPA-101200 locus appeared to inhibit expression of the QTL linked to UBC-333 [subscript 550]. ¶ To further analyse the quantitative trait loci (QTLs) controlling NESTUR, a linkage map was constructed from RAPD markers segregating in 93 haploid progeny of another full sib cross (30040 x 80121) (experiment 2). Two hundred and sixty-two (262) markers were mapped to 14 linkage groups of at least 7 markers, ranging in size from 39 to 183 cM. The 14 linkage groups covered approximately 1511 cM of genetic map distance. ¶ In experiment 3, the linkage map was used to map QTLs controlling NESTUR, as well as increments in seedling stem diameter, volume, and height and needle volume. Altogether, five putative QTLs were detected for NESTUR, with explained variation ranging from 9 to 22%. Of the five QTLs detected, 3 were coincidental with those for stem growth in height, diameter and volume. The two QTL positions that were unique to NESTUR were flanked by QTLs for the component traits. Together, effects of the five QTLs explained 48% of the total phenotypic variation for NESTUR. ¶ Ability of identified markers to predict the phenotype and seedlings with growth potential was assessed in the cross 30040 x 80121, using six RAPD markers associated with NESTUR at ANOVA P-levels of 0.01-0.001 (experiment 4). The correlation between observed NESTUR and predicted values was 0.70. Differences in observed vs. predicted values were not large and did not indicate serious misclassifications, such as classification of an upper ranking individual into the lower group, or vice versa. ¶ Over a two-year growth period, the ability of NESTUR to predict stem growth was strongly affected by seedling age. In contrast, markers linked to NESTUR showed a consistent ability to predict stem growth, irrespective of seedling age. Compared with the top 1% of the original population, seedlings selected for their genotypic values showed a higher stem volume growth of 103% in the first year, and 76% in the second year. ¶ The expression of QTLs for stem volume, stem diameter, height, number of branches, number of whorls, and branches/whorl were compared at 5, 12, and 24 months of age. Two QTLs detected for height showed contrasting expression over two years, one was gradually reduced from LOD of 2.70 to 0.43 and the other increased from 1.12 to 2.45. Compared with the pattern observed for height, LOD scan profiles for diameter and volume showed less temporal change of peaks, suggesting that the genetic control for height growth is probably more unstable than that of diameter. QTLs controlling the phenotype at the time of measurement (ie the final phenotype) showed similar magnitude of effects on that trait's respective increments (or growth rate).

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.

<p>The objective of the study was to conduct an internal validation on miniSTR systems and apply it to cases received from the South African Missing Persons Task Team (SAMPTT). This was prompted by the fact that miniSTR systems have been shown to out perform some of the commercial kits available in the time of the study and provide an alternative to mtDNA when analysing degraded DNA from skeletal remains and that the DNA extracted from skeletal remains received from the SAMPTT would be degraded due to the remains generally being fragmented or charred and buried for many years. The miniSTR loci chosen for validation comprised the Combined DNA Index System (CODIS) thirteen core loci and were arranged into four triplexes and one uniplex.</p>

No description available.