Doctor of Philosophy / Before the early 2000s, many producers were heavily selecting for production traits without accounting for the negative relationship between many production traits and fertility. The large decrease in fertility that occurred as a result of these selection practices put a heavy burden on producers. Replacement heifer development is the third largest cost incurred by producers, behind feed and labor. Consequently, the failure of a heifer to conceive in her first breeding season translates directly into financial loss for the producer and lasting consequences on the animal's longevity and performance in subsequent breeding seasons. To make improvements in a trait, or traits, of interest, producers often selectively breed two animals with desirable characteristics. However, the complex nature of fertility traits limits the effectiveness of this method. As a result, researchers have been attempting to identify biological molecules whose abundance differs between cattle of differing fertility potential, termed molecular markers, that could be used to identify superior cattle earlier and more accurately. While a good amount of research has been conducted in mature cows and in a laboratory setting, very few studies have attempted to identify molecular markers of fertility in heifers. Therefore, the objective of this work was to identify different biological molecules (genomic variants, genes, proteins, and metabolites) whose abundance differed between heifers with varying fertility potential. Investigation into DNA variations led to the identification of three variants that were associated with fertility, 16 variants that were associated with health, and 29 variants that were associated with both health and fertility. Given that some variants can impact a trait by changing gene expression, we attempted to identify variations in RNA that were having this effect on heifer fertility. Although some variants were found to influence gene expression, we were unable to correlate these changes with fertility differences. However, in a different study, we were able to identify two genes (APMAP and DNAI7), as well as one protein (alpha-ketoglutarate-dependent-dioxygenase FTO), that differed significantly between fertile and sub-fertile heifers. Importantly, the results of this study allowed us to create a biological profile that was capable of accurately distinguishing 21/22 heifers based on their fertility potential. Finally, investigation of the metabolite profile revealed one metabolite (2-dehydro-D-gluconate) that was differentially abundant between fertile and sub-fertile heifers. Overall, this work sheds light on the complex nature of heifer fertility and provides several potential molecular that could be used to distinguish between heifers of varying reproductive potential.

Kaukolės veidinės dalies anomalijos, ypač lūpos ir (arba) gomurio nesuaugimai (L/GN), yra vienos dažniausių žmogaus įgimtų ydų. Įvairių autorių duomenimis jų dažnumas svyruoja nuo 0,4 iki 2,0 1000 gimusiųjų. Sparčiai tobulėjant molekulinės genetikos metodams ir statistinės analizės įrankiams vis daugiau kandidatinių L/GN genetinių sričių identifikuojama plataus masto viso genomo tyrimais. Šio darbo tikslas buvo nustatyti genų kandidatų alelius, lemiančius polinkį (nesindrominiam) lūpos nesuaugimui su arba be gomurio nesuaugimo ir izoliuotam gomurio nesuaugimui Lietuvos pacientų grupėje, tiriant 42–jų genų galimai lemiančių L/GN genominius žymenis molekuliniais genetiniais metodais ir taikant statistinę asociacijos analizę nepusiausviro perdavimo testu. Taikant šeimų tyrimo ir atvejo – kontrolės tyrimo asociacijos analizės strategijas pacientų su lūpos nesuaugimu su arba be gomurio nesuaugimo grupėje, patvirtinti TIMP2, BMP2, FN1 genų kandidatų variantai lemiantys lūpos nesuaugimus su arba be gomurio nesuaugimo ir gomurio nesuaugimus Lietuvos populiacijoje, COL11A1, COL11A2, COL2A1 genų kandidatų variantai lemiantys gomurio nesuaugimus Lietuvos populiacijoje. Įvertinus Lietuvos, Latvijos ir Estijos pacientų su L/GN asociacijos analizės rezultatus, buvo nustatyta, kad Šiaurės Rytų Europos populiacijose polinkį lūpos nesuaugimui su arba be gomurio nesuaugimo lemia FGF1, TIMP2 genų variantai ir polinkį gomurio nesuaugimui COL2A1, COL11A2 genų variantai. Lūpos nesuaugimus su arba... [toliau žr. visą tekstą] / The incidence of cleft lip and (or) palate (CL/P) varies from 0.4 to 2.0 in 1000 live births across populations. More and more CL/P candidate loci are being confirmed using novel genome-wide methods of molecular genetics and tools of statistical analysis. The aim of this study was to identify the alleles of the candidate genes for cleft lip with or without cleft palate and isolated cleft palate in the Lithuanian patient group, applying the molecular genotyping of the genomic markers in 42 CL/P candidate genes and association analysis using transmission disequilibrium test approach. Using families’ studies and case – control studies approach CL/P candidate genes TIMP2, BMP2, FN1 variants were confirmed for cleft lip with or without cleft palate and COL11A1, COL11A2, COL2A1 for cleft palate only in the population of Lithuania. Estimating the association analysis results of the CL/P patients of Lithuania, Latvia and Estonia, FGF1, TIMP2 were identified as candidate genes for cleft lip with or without cleft palate and COL2A1, COL11A2 for isolated cleft palate in the North East European populations. Different genomic risk variants have influence for cleft lip with or without cleft palate and for cleft palate only, which confirms the hypothesis of different mechanisms for these two phenotypes.

The incidence of cleft lip and (or) palate (CL/P) varies from 0.4 to 2.0 in 1000 live births across populations. More and more CL/P candidate loci are being confirmed using novel genome-wide methods of molecular genetics and tools of statistical analysis. The aim of this study was to identify the alleles of the candidate genes for cleft lip with or without cleft palate and isolated cleft palate in the Lithuanian patient group, applying the molecular genotyping of the genomic markers in 42 CL/P candidate genes and association analysis using transmission disequilibrium test approach. Using families’ studies and case – control studies approach CL/P candidate genes TIMP2, BMP2, FN1 variants were confirmed for cleft lip with or without cleft palate and COL11A1, COL11A2, COL2A1 for cleft palate only in the population of Lithuania. Estimating the association analysis results of the CL/P patients of Lithuania, Latvia and Estonia, FGF1, TIMP2 were identified as candidate genes for cleft lip with or without cleft palate and COL2A1, COL11A2 for isolated cleft palate in the North East European populations. Different genomic risk variants have influence for cleft lip with or without cleft palate and for cleft palate only, which confirms the hypothesis of different mechanisms for these two phenotypes. / Kaukolės veidinės dalies anomalijos, ypač lūpos ir (arba) gomurio nesuaugimai (L/GN), yra vienos dažniausių žmogaus įgimtų ydų. Įvairių autorių duomenimis jų dažnumas svyruoja nuo 0,4 iki 2,0 1000 gimusiųjų. Sparčiai tobulėjant molekulinės genetikos metodams ir statistinės analizės įrankiams vis daugiau kandidatinių L/GN genetinių sričių identifikuojama plataus masto viso genomo tyrimais. Šio darbo tikslas buvo nustatyti genų kandidatų alelius, lemiančius polinkį (nesindrominiam) lūpos nesuaugimui su arba be gomurio nesuaugimo ir izoliuotam gomurio nesuaugimui Lietuvos pacientų grupėje, tiriant 42–jų genų galimai lemiančių L/GN genominius žymenis molekuliniais genetiniais metodais ir taikant statistinę asociacijos analizę nepusiausviro perdavimo testu. Taikant šeimų tyrimo ir atvejo – kontrolės tyrimo asociacijos analizės strategijas pacientų su lūpos nesuaugimu su arba be gomurio nesuaugimo grupėje, patvirtinti TIMP2, BMP2, FN1 genų kandidatų variantai lemiantys lūpos nesuaugimus su arba be gomurio nesuaugimo ir gomurio nesuaugimus Lietuvos populiacijoje, COL11A1, COL11A2, COL2A1 genų kandidatų variantai lemiantys gomurio nesuaugimus Lietuvos populiacijoje. Įvertinus Lietuvos, Latvijos ir Estijos pacientų su L/GN asociacijos analizės rezultatus, buvo nustatyta, kad Šiaurės Rytų Europos populiacijose polinkį lūpos nesuaugimui su arba be gomurio nesuaugimo lemia FGF1, TIMP2 genų variantai ir polinkį gomurio nesuaugimui COL2A1, COL11A2 genų variantai. Lūpos nesuaugimus su arba... [toliau žr. visą tekstą]

Die räumliche Organisation des Chromatins im Zellkern ist für die Regulierung der Genexpression von großer Bedeutung. Genomische Varianten können die räumliche Organisation jedoch stören und Fehlbildungen und Krankheiten verursachen. In diploiden Genomen sind die meisten genomischen Varianten heterozygot und beeinflussen hauptsächlich das homologe Chromosom, auf dem sie sich befinden. Daher ist eine allelspezifische Analyse wichtig, erweist sich aber mit aktuellen Methoden zur Erfassung der Chromatinkonformation als äußerst schwierig. Erstens ist der Haplotyp, der die Verteilung unterschiedlicher Allele über die homologen Chromosomen beschreibt, oft unbekannt. Zweitens ist, insbesondere in Genomen mit geringer Variantendichte, wie dem menschlichen Genom, eine eindeutige Zuordnung der sequenzierten Genomabschnitte (Reads) zu ihrem Ursprungschromosom häufig nicht möglich, was die Erstellung haplotypspezifischer Chromatinkontaktmatrizen von guter Qualität verhindert. Genome Architecture Mapping (GAM) ist eine vielversprechende neue Methode mit dem Potential zur haplotypspezifischen Analyse der Chromatinkonformation. In dieser Dissertation zeige ich zunächst, dass GAM-Daten wertvolle Haplotypinformationen enthalten. Dann stelle ich GAMIBHEAR vor, einen graphenbasierten Ansatz, der die von GAM-Daten abgeleiteten Phaseninformationen nutzt, um genaue und vollständige Haplotypen zu rekonstruieren. Schließlich stelle ich Co-Phasing vor, eine neue Read-Phasing-Strategie, die erstmalig die eindeutige Zuordnung von variantenfreien Reads zu ihrem homologen Ursprungschromosom ermöglicht und somit auch die Erstellung detaillierter haplotypspezifischer Chromatinkontaktmatrizen in Maus und Mensch. Im Gegensatz zu früheren Erkenntnissen belegen meine Ergebnisse große Unterschiede in der räumlichen Organisation homologer Chromosomenkopien und ermöglichen erstmals einen sehr detaillierten Einblick in die haplotypspezifische Chromatinkonformation des menschlichen Genoms. / The spatial organization of chromatin in the nucleus plays an essential role in precise gene expression. Genomic variants can disrupt this spatial organization, potentially causing malformations and diseases. In diploid genomes, most genomic variants are heterozygous and mainly influence the homologous chromosome they reside on. Studying the effects of these variants in an allele-specific manner is crucial but has proven challenging using current state-of-the-art techniques. First, the haplotype describing the distribution of variant alleles over the homologous chromosomes is often unknown. Second, especially in genomes with a low variant density, such as the human genome, most sequencing reads map to genomic regions that are identical between homologous chromosomes, making it difficult to determine their origin. Thus, the read-phasing efficiency is insufficient to generate haplotype-specific chromatin contact matrices of good quality. Genome Architecture Mapping (GAM) is a promising new method for haplotype-specific analysis of chromatin conformation. In this thesis, I first demonstrate the ability of GAM data to provide valuable haplotype information. Then, I introduce GAMIBHEAR, a graph-based approach that leverages the GAM-derived phase information to infer accurate and complete haplotypes. Finally, building on GAMIBHEAR, I present Co-Phasing, a novel read-phasing strategy that allows for the unique assignment of variant-free reads to their homologous chromosome of origin and thus enables the creation of detailed haplotype-specific chromatin contact matrices in mouse and human. In contrast to previous findings, my results show significant differences in the spatial organization of homologous chromosomes and provide the first detailed view of haplotype-specific chromatin conformation in the human genome.