Growth rate and flowering time are agriculturally important traits that are linked to fitness, productivity and reproductive success of plants. To study the genetic basis for natural variation in growth rate and flowering time between local accessions of Arabidopsis thaliana, hybrids were produced between fast growing / late flowering and slow growing / early flowering parents. F3 and F5 hybrid families were grown under a range of conditions – under a constant controlled environment, outside over the winter and outside in spring and early summer. Growth rates were estimated from repeated images of rosettes. Flowering time, as number of leaves to flower, was also recorded both in control and natural conditions for F5 lines. Damage by slugs and stress-induced production of anthocyanin pigments were also recorded for plants grown outside. Broad-sense heritability estimates were higher for F5 families than F3, in which more loci will segregate, and ranged from 48% to 89%. No significant correlation between growth rates under different environments was observed in most cases for F3 populations, however significant correlations were detected for F5 families outside and under controlled conditions, suggesting that same genes can affect growth rate in more than one environment. The genotypes of F3 families were determined at thirty-nine SSLP (simple sequence length polymorphism) loci and used in regression with phenotype data to search for quantitative trait loci (QTL). Significant QTLs were detected in F3 families for growth rate, flowering time and anthocyanin production, but not for herbivore damage. To confirm QTL detected in the F3 and to detect additional loci, bulk segregant analysis was carried out in F5 families grown under different conditions. Potentially linked markers were tested further in individual F5 plants and QTL mapped on a finer scale in F5 families that remained heterozygous for candidate regions. VIP5 and LDL1 were selected as potential candidate genes for flowering time variation. These genes were sequenced for two parental alleles. A transposon insertion and 5’ UTR deletion were found in the LDL1 allele from the late flowering parent and SNPs (single nucleotide polymorphisms) were observed throughout the gene. However both alleles appeared to be expressed at similar levels. Transgenic lines have been produced carrying the LDL1 allele from the early flowering parent (4D1) in the background of the later flowering parent (11C1). This work is on-going and will hopefully reveal whether LDL1 underlies differences in flowering behaviour seen between 11C1 and 4D1.

Thesis (PhD (Genetics))--Stellenbosch University, 2010. / ENGLISH ABSTRACT: The South African abalone, Haliotis midae, is a commercially valuable mollusc and is mostly exported to the Far East. Genetics research on H. midae has increased substantially since a genetic improvement programme was introduced in 2006 by collaboration between Stellenbosch University, government and industry partners. The development of molecular markers, QTL-mapping, gene-expression and genome manipulations are the main focuses of the research currently being conducted. The end goal is to create high quality and fast growing animals for the industry. The present study focused on the development of microsatellite markers and the detection of quantitative trait loci (QTL) affecting growth traits (shell length, shell width, wet weight) in this species. A combination of three methods, namely selective genotyping and bulked segregant analysis (pooling analysis), single marker regression and interval mapping were used to identify putative QTL in two full-sib families from two different farmed locations. Additional methods and protocols were developed that can assist the industry in other molecular research aspects. A total of 125 microsatellite loci were characterised. A total of 82 of these loci were isolated using second generation sequencing, a first for any abalone species. A preliminary, low-density framework linkage map was constructed containing 50 loci that mapped to 18 linkage groups. The observed genome length was 148.72cm with coverage of ±47%. QTL analyses revealed two putative QTL for shell width and wet weight, with 17% and 15% variance explained, that mapped on one linkage group in the first family and three putative QTL, for shell length, shell width and wet weight, with 33%, 28.5% and 31.5% variance explained, that mapped on one linkage group in the second family. Additional methods and protocols developed include an automated high-throughput DNA isolation protocol, a real-time PCR assay for H. midae x H. spadicea hybrid verification, a triploid verification microsatellite assay and a pre- and post-PCR multiplex setup and optimisation protocol. Future studies focussing on QTL and marker assisted selection (MAS) should verify the QTL found in this study and also utilise additional family structures and determine QTL-marker phase within the commercial populations. / AFRIKAANSE OPSOMMING: Die Suid-Afrikaanse perlemoen, Haliotis midae, is ’n kommersieel waardevolle weekdier en word hoofsaaklik na die Verre-Ooste uitgevoer. Genetiese navorsing op H. midae het aansienlik toegeneem sedert ’n genetiese verbeteringsprogram in 2006 deur samewerking tussen die Universiteit van Stellenbosch, die regering en industrievennote ingebring is. Die ontwikkeling van molekulêre merkers, KEL-kartering, geen-uitdrukking en genoom manipulasies is die hooffokusse van die navorsing wat tans uitgevoer word. Die einddoel is om hoë kwaliteit en snelgroeiende diere vir die industrie te skep. Die huidige studie het op die ontwikkeling van mikrosatelliet merkers en die opsporing van groeiverwante (skulplengte, -breedte en nat gewig) kwantitatiewe eienskap lokusse (KEL) in hierdie spesie gefokus. ’n Kombinasie van drie metodes, naamlik selektiewe genotipering en versamelde segregaat analise (samevoegingsanalise), enkel merker regressie en intervalkartering is gebruik om waarskynlike KEL in twee vol-sibbe families van twee verskillende produksiegebiede te identifiseer. Aanvullende metodes en protokolle is ontwikkel wat die industrie in ander molekulêre navorsingsaspekte kan ondersteun. ’n Totaal van 125 mikrosatelliet lokusse is beskryf. ’n Totaal van 82 van hierdie lokusse is deur die gebruik van derde generasie volgordebepaling geïsoleer, ’n eerste vir enige perlemoen spesie. ’n Voorlopige, laedigtheid raamwerkkoppelingskaart is saamgestel met 50 lokusse wat op 18 koppelingsgroepe gekarteer is. Die waarneembare genoomlengte was 148.72cm met ’n dekking van ±47%. KEL-analises het twee waarskynlike KEL vir skulpbreedte en nat gewig blootgelê wat 17% en 15% variasie verduidelik en is op een koppelingsgroep in die eerste familie gekarteer asook drie waarskynlike KEL, vir skulplengte, -breedte en nat gewig wat 33%, 28.5% en 31.5% variasie verduidelik en is op een koppelingsgroep in die tweede familie gekarteer. Aanvullende metodes en protokolle wat ontwikkel is, sluit ’n geoutomatiseerde hoë-deurgang DNS-isolasieprotokol, ’n intydse PKR-proef vir H. midae x H. spadicea hibried verifikasie, ’n triploïed verifikasie mikrosatellietproef en veelsoortige pre- en post-PKR opstelling en optimaliseringsprotokol in. Toekomstige studies wat fokus op KEL en merker ondersteunde seleksie (MOS) behoort die KEL wat in hierdie studie gevind is te verifieer en ook bykomende familie strukture te benut om KEL-merker fases binne die kommersiële populasie te bepaal.

It is well documented that both quantitative and qualitative changes in the murine hematopoietic stem cell (HSC) population occur with age. In mice, the effect of aging on stem cells is highly strain-specific, thus suggesting genetic regulation plays a role in HSC aging. In C57BL/6 (B6) mice, the HSC population steadily increases with age, whereas in DBA/2 (D2) mice, this population declines. Our lab has previously mapped a quantitative trait locus (QTL) to murine chromosome 2 that is associated with the variation in frequency of HSCs between aged B6 and D2 mice. In these dissertation studies, I first aim to characterize the congenic mouse model which was generated by introgressing D2 alleles in the QTL onto a B6 background. Using a surrogate assay to mimic aging, I analyzed the cell cycle, apoptotic and self-renewal capabilities of congenic and B6 HSCs and show that D2 alleles in the QTL affect the apoptotic and selfrenewal capabilities of HSCs. In the second aim of these studies, I used oligonucleotide arrays to compare the differential expression of B6 and congenic cells using a population enriched for primitive stem and progenitor cells. Extensive analysis of the expression arrays pointed to two strong candidates, the genes encoding Retinoblastoma like protein 1 (p107) and Sorting nexin 5 (Snx5). B6 alleles were associated with increased p107 and Snx5 expression in old HSCs therefore both genes were hypothesized to be positive regulators of stem cell number in aged mice. Finally, in the third aim of these studies, I show that the individual overexpression of p107 and Snx5 in congeic HSCs increases day35 cobblestone area forming cell (CAFC) numbers, therefore confirming their roles as positive regulators of HSC number in vitro. These studies uncover novel roles for p107 and Snx5 in the regulation of HSC numbers and provide additional clues in the complex regulation of HSC aging.

Identification of allelic variation through quantitative trait loci (QTL) mapping offers possibilities for the improvement of quantitatively inherited traits. This requires a genetic map along with the phenotypic characterization of a mapping population. A doubled haploid (DH) Polo X Topas population consisting of 194 lines and a recombinant inbred line population of 92 lines was developed. Individual genetic maps derived from each population were integrated into a consensus map. The DH-based genetic map was used for QTL mapping. The DH-based map was comprised of 620 loci that were assembled into 19 linkage groups that were anchored to the B. napus chromosomes. The DH-based map covered 2244.1 cM genomic distance with an average marker interval of 3.7 cM. The DH population was phenotyped in four environments with each line replicated twice in a randomized complete block design. Days to flowering was recorded and oil content and fatty acid composition were determined using Near Infrared spectroscopy (NIR) and Gas Chromatography, respectively. Fourteen QTL were identified for oil content, 33 QTL for palmitic acid content, 18 QTL for stearic acid content, 21 QTL for oleic acid content, 20 QTL for linoleic acid content, 23 QTL for linolenic acid content, 16 QTL for arachidic acid content and 14 QTL for flowering time. Oil content QTL were identified on five linkage groups, A3, A10, C1, C5, and C6. An oil content QTL, qOIL-A10c appeared in all four environments, whereas qOIL-A10a appeared in only one environment but explained 26.99% variation. The oil content in the population ranged from 35% to 55.5% with the parents having values of 42% to 46%. Two genomic regions on C3, with map positions at 147.83 cM and 154.55 cM harbored QTL (rQTL) for all the fatty acids studied. The additive effects of the rQTL reveal a correlation pattern which is supported by the phenotypic correlation observed between the fatty acids. This suggests rQTL have role in the fatty acid composition and possibly determine total seed oil content. The rQTL and flanking markers of the identified QTL offer utility in further development of B. napus. / October 2015

Par définition le déséquilibre de liaison (Linkage Disequilibrium, LD) décrit les associations préférentielles entre allèles de deux locus. Ce concept est devenu un outil indispensable pour la cartographie fine de locus quantitatifs (QTL), par l'identification de déséquilibres d'associations entre allèles à un locus marqueur (ou à un ensemble de locus marqueurs) et à un locus impliqué dans la variation d'un caractère quantitatif. La création et l'intensité du LD sont dépendantes des forces évolutives qui ont construit la population. Parmi ces forces, la dérive génétique et la sélection sont particulièrement actives dans les populations d'animaux de rente. Cette thèse a pour but d'étudier l'influence de la sélection sur la structure du déséquilibre de liaison autour d'un locus quantitatif, ainsi que son impact sur la précision de cartographie fine des QTL. Un logiciel de simulation de populations a été développé dans le cadre de la thèse. A partir d'une population en équilibre de liaison, il permet de générer du LD dans des générations dites historiques, grâce à différentes forces évolutives. La détection de QTL est appliquée aux générations suivantes, de généalogie connue. Pour ces dernières générations, les principaux dispositifs de détection de QTL de génétique animale sont décrits dans le simulateur. Les données exploitées dans cette thèse sont issues de ce logiciel. Le LD a été mesuré par le D' et le

This thesis addresses the dissection and characterisation of quantitative trait loci (QTL) affecting production traits in sheep. Firstly, the association between specific genetic polymorphisms and complex variation in weight, muscle and fat depositions was investigated. Research concentrated on assessing the presence, correspondence and significance of two single nucleotide polymorphisms (SNPs) in the GDF8 region of ovine chromosome 2, reportedly affecting muscle production. Commercial populations of British Texel, Suffolk and Charollais sheep were studied. The SNPs were absent in Suffolk and almost fixed in Texel breeds. In the Charollais population, the SNPs segregated at intermediate frequencies and a significant association was found between these polymorphisms and muscle depth. The previously proposed causative allele at one of the loci resulted in increased muscle depth and, at allele frequency of 0.5, this locus would explain one third of the additive genetic variance for the trait. Partial recessive allelic expression is proposed by genotypic value predictions and is consistent with the previously postulated molecular mechanism by which it gives rise to muscle changes. Secondly, the thesis focused on detection of QTL associated with growth. Live weight is a composite of growth rates over time, with inter-age genetic correlations for live weight decreasing as time between weight measurements increases. To explore whether observed genetic correlation patterns translate into distinct loci acting on weight at different growth stages, a novel method was developed and the applicability of a second proposed method was explored. Both methods allowed simultaneous analysis of multiple live weights per animal, while accounting differently for the correlation among measurements ordered in time. In the first approach, a growth curve technique was developed and employed to map growth QTL for curve parameters and predicted growth descriptors. A study of actual live weights identified significant QTL at different ages on distinct chromosomes, with QTL significance and variance changing over time. Further application of this technique on a simulated dataset validated its effectiveness in detecting age-dependent QTL. An extension of the procedure resulted in a novel technique for genomic evaluation of longitudinal traits. In the second method examined, random regression (RR) models were applied for dissection of growth QTL. Systematic model selection and inclusion of relevant random effects resulted in apparently significant QTL, but the method was computationally demanding, model choice proved challenging and the results were questioned. To further explore the method, RR models were applied to various simulated growth phenotypes composed of time-dependent QTL trajectories, polygenic and environmental effects. Statistically optimal RR models succeeded in identifying significant QTL and predicting the simulated time-dependence for most scenarios. However, the issue of model choice was again prominent, as suboptimal models resulted in unreliable QTL variance trajectories and pronounced confounding between different time-dependent effects. Thus, the growth curve approach appeared to be the more flexible and robust process for analysing longitudinal data to map agedependent QTL.

The virulence of Plasmodium falciparum is largely attributed to the ability of asexual blood-stage parasites to cytoadhere to the microvascular endothelium of the human host. This pathogenic behavior is mediated by the primary parasite virulence factor P. falciparum erythrocyte membrane protein 1 (PfEMP1), an understanding of which is crucial to develop interventions to ameliorate the morbidity and mortality of P. falciparum malaria. The work presented in this thesis describes the application of a phenotype-to-genotype experimental approach to identify novel parasite proteins involved in the trafficking and display of PfEMP1. Guided by the overall hypothesis that the in vitro culture-adapted parasite line 3D7 harbors 1 or more genetic determinants of impaired PfEMP1 trafficking, surface PfEMP1 levels were first measured in 3D7, the presumably trafficking-competent parasite line HB3, and 16 unique progeny from an HB3 x 3D7 genetic cross (chapter 2). These phenotypes were then combined with genome-wide SNP data in QTL analysis to identify genetic polymorphisms potentially responsible for the impaired trafficking in 3D7 (chapter 3). A near-significant QTL containing a single protein-coding gene, the putative kinesin Pf3D7_1245600, was identified, characterized, and investigated in CRISPR-Cas9-driven allele-exchange parasite transfection experiments to establish a causal link between the gene and PfEMP1 trafficking (chapter 4). The parasite transfections were unsuccessful, but the potential role of Pf3D7_1245600 in PfEMP1 trafficking was indirectly assessed through the disruption of microtubules with colchicine (chapter 4), which significantly impacted the surface PfEMP1 levels of HB3 but not 3D7. The findings of this thesis suggest that kinesins and microtubules may play previously unconsidered roles in the regulation, production, or trafficking of PfEMP1.

Advancing the production efficiency and profitability of aquaculture is dependent upon the ability to utilize a diverse array of genetic resources. The ultimate goals of aquaculture genomics, genetics and breeding research are to enhance aquaculture production efficiency, sustainability, product quality, and profitability in support of the commercial sector and for the benefit of consumers. In order to achieve these goals, it is important to understand the genomic structure and organization of aquaculture species, and their genomic and phenomic variations, as well as the genetic basis of traits and their interrelationships. In addition, it is also important to understand the mechanisms of regulation and evolutionary conservation at the levels of genome, transcriptome, proteome, epigenome, and systems biology. With genomic information and information between the genomes and phenomes, technologies for marker/causal mutation-assisted selection, genome selection, and genome editing can be developed for applications in aquaculture. A set of genomic tools and resources must be made available including reference genome sequences and their annotations (including coding and non-coding regulatory elements), genome-wide polymorphic markers, efficient genotyping platforms, high-density and high-resolution linkage maps, and transcriptome resources including non-coding transcripts. Genomic and genetic control of important performance and production traits, such as disease resistance, feed conversion efficiency, growth rate, processing yield, behaviour, reproductive characteristics, and tolerance to environmental stressors like low dissolved oxygen, high or low water temperature and salinity, must be understood. QTL need to be identified, validated across strains, lines and populations, and their mechanisms of control understood. Causal gene(s) need to be identified. Genetic and epigenetic regulation of important aquaculture traits need to be determined, and technologies for marker-assisted selection, causal gene/mutation-assisted selection, genome selection, and genome editing using CRISPR and other technologies must be developed, demonstrated with applicability, and application to aquaculture industries. Major progress has been made in aquaculture genomics for dozens of fish and shellfish species including the development of genetic linkage maps, physical maps, microarrays, single nucleotide polymorphism (SNP) arrays, transcriptome databases and various stages of genome reference sequences. This paper provides a general review of the current status, challenges and future research needs of aquaculture genomics, genetics, and breeding, with a focus on major aquaculture species in the United States: catfish, rainbow trout, Atlantic salmon, tilapia, striped bass, oysters, and shrimp. While the overall research priorities and the practical goals are similar across various aquaculture species, the current status in each species should dictate the next priority areas within the species. This paper is an output of the USDA Workshop for Aquaculture Genomics, Genetics, and Breeding held in late March 2016 in Auburn, Alabama, with participants from all parts of the United States.

The goal of this project was to determine the genetic architecture of water-use efficiency (WUE) for foxtail pine, which included genomic loci, and effect sizes of this trait. Foxtail pine is a subalpine endemic conifer that inhabits two distinct regional populations separated by 500 km in the mountains of California. In order to achieve this goal, a robust linkage map containing thousands of genetic markers was created using four megagametophyte arrays ranging in size from approximately 70 to 95 megagametophytes. Quantitative trait loci (QTL) discovered for WUE were mapped along the linkage map using linear mixed models and five half-sibling families grown in a common garden. Effect sizes of these QTL were tested for differences between the two regional populations of foxtail pine.

The PRDM9 (PR domain containing 9) protein is an epigenetic factor that trimethylates lysine 4 of histone H3 and thereby determines the future meiotic double-strand breaks - sites important for proper segregation of homologous chromosomes. Males of the Mus musculus domesticus (Mmd) origin with homozygous deletion in Prdm9 (Prdm9-/- ) are sterile with a complete arrest in meiotic prophase I, in contrast to the same mutant males of the M. m. musculus (Mmm) subspecies. The aim of this diploma thesis was to identify the genomic loci responsible for the phenotypic difference of these Prdm9-/- males. The major research tool was a population of 182 Mmm x Mmd Prdm9-/- males. The mapping method of quantitative trait loci (QTLs) was based on relating the genotypes of single-nucleotide and microsatellite polymorphisms to the observed phenotypes. At least two QTLs on Chr X were identified. The Mmm alleles of these QTLs reduced fertility of Prdm9-/- males. Both QTLs were confirmed and narrowed down using two types of subconsomic strains. It was not possible to confirm other QTLs, particularly on autosomes. This QTL mapping is the first step towards the identification of genes that modify the resulting phenotype of Prdm9-/- animals. This identification should help designing studies of human infertility that...