The programmable network envisioned in the 1990s within standardization and research for the Intelligent Network is currently coming into reality using IPbased Next Generation Networks (NGN) and applying Service-Oriented Architecture (SOA) principles for service creation, execution, and hosting. SOA is the foundation for both next-generation telecommunications and middleware architectures, which are rapidly converging on top of commodity transport services. Services such as triple/quadruple play, multimedia messaging, and presence are enabled by the emerging service-oriented IPMultimedia Subsystem (IMS), and allow telecommunications service providers to maintain, if not improve, their position in the marketplace. SOA becomes the de facto standard in next-generation middleware systems as the system model of choice to interconnect service consumers and providers within and between enterprises. We leverage previous research activities in overlay networking technologies along with recent advances in network abstraction, service exposure, and service creation to develop a paradigm for a service environment providing converged Internet and Telecommunications services that we call Service Broker. Such a Service Broker provides mechanisms to combine and mediate between different service paradigms from the two domains Internet/WWW and telecommunications. Furthermore, it enables the composition of services across these domains and is capable of defining and applying temporal constraints during creation and execution time. By adding network-awareness into the service fabric, such a Service Broker may also act as a next generation network-to-service element allowing the composition of crossdomain and cross-layer network and service resources. The contribution of this research is threefold: first, we analyze and classify principles and technologies from Information Technologies (IT) and telecommunications to identify and discuss issues allowing cross-domain composition in a converging service layer. Second, we discuss service composition methods allowing the creation of converged services on an abstract level; in particular, we present a formalized method for model-checking of such compositions. Finally, we propose a Service Broker architecture converging Internet and Telecom services. This environment enables cross-domain feature interaction in services through formalized obligation policies acting as constraints during service discovery, creation, and execution time. / Das programmierbare Netz, das Ende des 20. Jahrhunderts in der Standardisierung und Forschung für das Intelligente Netz entworfen wurde, wird nun Realität in einem auf das Internet Protokoll basierendem Netz der nächsten Generation (Next Generation Network). Hierfür kommen Prinzipien aus der Informationstechnologie, insbesondere aus dem Bereich dienstorientierte Architekturen (Service-Oriented Architecture / SOA) für die Diensterstellung, -ausführung und -betrieb zum Tragen. SOA bietet hierbei die theoretische Grundlage für Telekommunikationsnetze, vor allem jedoch für die dazugehörigen Dienstplattformen. Diese erlauben dem Telekommunikationsbetreiber seine Position in einem offenen Marktplatz der Dienste auszubauen. Dazu bedarf es allerdings möglichst flexibler Dienstumgebungen, die die Kooperation zwischen Dienstanbietern und Nutzern aus unterschiedlichsten Domänen durch Unterstützung geeigneter Werkzeuge und Mechanismen fördert. Im Rahmen dieser Dissertation definieren wir aufbauend auf Forschungsergebnisse im Bereich Overlay-Netze, Netzabstraktion und Zugriff auf exponierte Dienste eine Service Broker genannte Dienstumgebung für konvergente Internet- und Telekommunikationsdienste. Dieser Service Broker stellt Mechanismen für die Komposition von Diensten und Mediation zwischen unterschiedlichen Dienstparadigmen und Domänenspezifika beim Dienstaufruf zur Verfügung. Der Forschungsbeitrag dieser Arbeit findet auf unterschiedlichen Ebenen statt: Aufbauend auf einer Analyse und Klassifikation von Technologien und Paradigmen aus den Bereichen Informationstechnologie (IT) und Telekommunikation diskutieren wir die Problemstellung der Kooperation von Diensten und deren Komposition über Domänengrenzen hinweg. In einem zweiten Schritt diskutieren wir Methoden der Dienstkomposition und präsentieren eine formalisierte Methode der modellbasierten Diensterstellung. Der Schwerpunkt der Arbeit liegt auf der Spezifikation der Service Broker Dienstumgebung und einem zugrundeliegenden Informations- und Datenmodell. Diese Architektur erlaubt die Komposition und Kooperation von Diensten über Domänengrenzen hinweg, um konvergente Internet- und Telekommunikationsdienste zu realisieren. Hierfür wird ein auf Obligationspolitiken basierendes Regelsystemformalisiert, das Interaktionen zwischen Dienstmerkmalen während der Diensterstellung und -ausführung definiert.

Fibre deployment of next-generation high-speed broadband networks is considered to be a decisive development for any information-based society, yet investment activities and especially the adoption of fibre-based broadband services take place only very gradually in most countries. This work identifies the most important determinants of the adoption of fibre-based broadband services, using the most recent panel data from the European Union member states (EU27) for the years from 2004 to 2012. The results show that the stricter previous broadband access regulation has a negative impact on adoption, while competitive pressure from mobile networks affects adoption in a non-linear manner. It appears that the approach of strict cost-based access regulation embedded in the EU regulatory framework is at odds with the targets outlined in the European Commission's "Digital Agenda". Finally, we also find strong evidence for network effects underlying the adoption process. (author's abstract) / Series: Working Papers / Research Institute for Regulatory Economics

Although fibre-deployment of next generation access (NGA) broadband networks is considered as a decisive development for any information-based society, investment activities and especially the adoption of fiber-based broadband services take place only very gradually in most countries. This work identifies the most important determinants of NGA broadband adoption, using most recent panel data from the European Union member states (EU27) for the years from 2004 to 2012. The results show that stricter previous broadband access regulation has a negative impact on NGA adoption, while competitive pressure from mobile networks affects NGA adoption in a non-linear manner. It appears that the approach of strict cost-based access regulation embedded in the EU regulatory framework is at odds with the ambitious targets outlined in the European Commission´s "Digital Agenda". Finally, we find strong evidence for network effects underlying the NGA adoption process. (author's abstract) / Series: Working Papers / Research Institute for Regulatory Economics

Fibre deployment of next-generation high-speed broadband networks is considered to be a decisive development for any information-based society, yet investment activities and especially the adoption of fibre-based broadband services take place only very gradually in most countries. This work identifies the most important determinants of the adoption of fibre-based broadband services, using the most recent panel data from the European Union member states (EU27) for the years from 2004 to 2012. The results show that the stricter previous broadband access regulation has a negative impact on adoption, while competitive pressure from mobile networks affects adoption in a non-linear manner. It appears that the approach of strict cost-based access regulation embedded in the EU regulatory framework is at odds with the targets outlined in the European Commission's "Digital Agenda". Finally, we also find strong evidence for network effects underlying the adoption process. (author's abstract) / Series: Working Papers / Research Institute for Regulatory Economics

This master's thesis deals with the creation of a tool for the extraction of methylation level from transposon sequences. Transposons are DNA elements with ability to move or copy themselves and their activity is regulated by DNA methylation. Sequence methylation information is stored in the bisulfite data and their processing is done with parts of two existing tools in a combination with implemented modules. Created tool takes into consideration unique challenges brought in the methylation calling process by transposable elements and it's functionality is presented on a set of experiments with simulated and real data.

Noroviruses are the major cause of acute viral gastroenteritis worldwide. Thus, rapid and reliable pathogen detection and control are crucial to avoid epidemic outbreaks. Peptides which bind to these viruses with high specificity and affinity could serve as small and stable recognition elements in biosensing applications for a point-of-care diagnostic of noroviruses. They can be identified by screening large phage display libraries using the biopanning technique. In the present study, this method was applied to identify norovirus-binding peptide motifs. For this purpose, a biopanning based on column chromatography was established, and three rounds of selections were performed. After the second round, the cosmix-plexing recombination technique was implemented to enhance the chance of obtaining peptides with very high affinity. Biopanning data evaluation was based on next-generation sequencing (NGS), to show that this innovative method can enable a detailed analysis of the complete sequence spectrum obtained during and after biopanning. Highly enriched motifs could be characterized by their large proportion of the amino acids W, K, R, N, and F. Neighbourhood analysis was exemplarily performed for selected motifs, showing that the motifs FAT, RWN, and KWF possessed the fingerprints with the largest differences relative to the original library. This thesis thus presents next-generation sequencing-based analysis tools, which could now be transferred to any other biopanning project. The identified peptide motifs represent promising candidates for a future examination of their norovirus-specific binding. A new option for testing such phage-target interactions in the context of biopanning selections was studied in the second part of the thesis. For this purpose, a phage-based microarray was developed as a miniaturized binding assay. As a prerequisite, the different immobilization behaviour of phages on positively and negatively charged surfaces was studied, and a non-contact printing technique for bacteriophages was developed. Subsequently, the interaction of phages and antibodies directed against phage coat proteins was characterized in enzyme-linked immunosorbent assays, and the protocol was successfully transferred to the non-contact printed phage spots. At the proof-of-concept level, the phage array could finally be integrated into a microfluidic setup, showing a higher signal-to-background ratio relative to the static phage array. These results point the way towards a microfluidic phage array, allowing online monitoring, automation, and parallelisation of the phage array analysis. / Noroviren gelten als Hauptursache akuter viraler Magen-Darm-Erkrankungen. Nur eine zeitnahe und verlässliche Detektion und Kontrolle dieser Pathogene kann epidemische Ausbrüche vermeiden. Um dies zu ermöglichen, könnten Peptide, die an diese Viren mit hoher Spezifität und Affinität binden, als kleine und stabile Erkennungselemente in biosensorischen Anwendungen eingesetzt werden. Solche Peptide können mithilfe der Biopanning-Technik identifiziert werden, die auf dem Screening großer Phagen-Display-Bibliotheken beruht. In der vorliegenden Arbeit wurde diese Methode genutzt, um Norovirus-bindende Peptidmotive zu identifizieren. Dazu wurde ein auf Säulenchromatographie basierendes Biopanning entwickelt und drei Selektionsrunden durchgeführt. Die Cosmix-Plexing-Rekombinationstechnik wurde nach der zweiten Runde eingesetzt, um die Wahrscheinlichkeit der Gewinnung hochaffiner Binder zu erhöhen. Die Auswertung der Biopanningdaten erfolgte mittels Hochdurchsatzsequenzierung (Next-Generation Sequencing). Es konnte gezeigt werden, dass diese innovative Methode die detailierte Analyse des kompletten Sequenzspektrums während und nach dem Biopanning ermöglicht. Stark angereicherte Motive konnten durch ihren hohen Anteil an den Aminosäuren W, K, R, N und F charakterisiert werden. Eine Nachbarschaftsanalyse wurde exemplarisch für ausgewählte Motive durchgeführt. Dabei wurden die stärksten Unterschiede im Fingerprint im Vergleich zur Ausgangsbibliothek bei den Motiven FAT, RWN und KWF gefunden. Diese Dissertation stellt damit auf Next-Generation Sequencing basierende Analysetechniken vor, die für weitere Biopanningprojekte übernommen werden können. Die identifizierten Peptidmotive könnten als vielversprechende Kandidaten zukünftig auf ihre Norovirus-spezifische Bindung hin getestet werden. Eine neue Möglichkeit, solche Phagen-Analyt-Interaktionen zu untersuchen, wurde im zweiten Teil der Dissertation untersucht. Dafür wurde als miniaturisierter Bindungsassay ein Phagen-basiertes Mikroarray entwickelt. Als Voraussetzung wurde zunächst das unterschiedliche Immobilisierungsverhalten von Bakteriophagen auf positiv und negativ geladenen Oberflächen untersucht und eine kontaktfreie Drucktechnik für Bakteriophagen etabliert. Anschließend wurde die Interaktion von Phagen und gegen sie gerichteten Antikörpern in Enzym-gekoppelten Immunadsorptionstests charakterisiert und das Protokoll erfolgreich auf die kontaktfrei gedruckten Phagenspots übertragen. Schließlich wurde erstmals die grundsätzliche Möglichkeit gezeigt, das Array in ein mikrofluidisches Setup zu integrieren, was zu einem höheren Signal-zu-Hintergrund-Verhältnis im Vergleich zum statischen Array führte. Diese Ergebnisse zeigen damit den Weg zu einem mikrofluidischen Phagen-Array auf, das sowohl die Möglichkeit des Online-Monitorings als auch der Automatisierung und Parallelisierung der Phagen-Array-Analyse bietet.

(English) Meiotic homologous recombination, homologous chromosomes synapsis, and F1 hybrid sterility (enabling formation of species) are mutually interconnected phenomenons, one being the prerequisite to the latter. In the present thesis, these phenomenons were investigated on a genetic and mechanistic level using a mouse subspecies as a model. Noncrossovers (NCOs, gene conversions), 90% prevalent resolution of Prdm9- determined meiotic double-strand breaks (DSBs), were uniquely identified and characterized on a chromosome-wide level. The mean gene conversion tract length, based on 94 NCOs events, was calculated to be 32 bp. On a local level, the NCOs overlapped the known hotspots of PRDM9-controlled histone trimethylation and DSB formation, indicating their origin in the standard meiotic DSB repair pathway. On chromosome-wide level, NCO and CO distributions differed, in particular COs being relatively preferred over NCOs in subtelomeric regions. A specific subset of nonparental/asymmetric NCOs and COs was underrepresented in our datasets, proposing their problematic repair, hypothetically enabled by sister chromatids, and thus not contributing to indispensable homologous synapsis. Genome-wide crossover (CO) rates, genetically and mechanistically crucial ~10% of DSB repair, were proven to be...

Analyzing the data produced by next-generation sequencing technologies relies on access to information synthesized based on previous research findings. The volume of data available in the literature is growing rapidly, and it is becoming increasingly necessary for researchers to use AI or other statistics-based approaches in the analysis of their datasets. In this project, knowledge graphs are explored as a tool for providing access to contextual gene-information available in scientific literature. The explorative method described in this thesis is based on the implementation and comparison of two approaches for knowledge graph construction, a rule-based statistical as well as a neural-network and co-occurrence based approach, -based on specific literature contexts. The results are presented both in the form of a quantitative comparison between approaches as well as in the form of a qualitative expert evaluation of the quantitative result. The quantitative comparison suggested that contrasting knowledge graphs constructed based on different approaches can provide valuable information for the interpretation and contextualization of key genes. It also demonstrated the limitations of some approaches e.g. in terms of scalability as well as the volume and type of information that can be extracted. The result further suggested that metrics based on the overlap of nodes and edges, as well as metrics that leverage the global topology of graphs are valuable for representing and comparing contextual information between knowledge graphs. The result based on the qualitative expert evaluation demonstrated that literature-derived knowledge graphs of gene-information can be valuable tools for identifying research biases related to genes and also shed light on the challenges related to biological entity normalization in the context of knowledge graph development. In light of these findings, automatic knowledge-graph construction presents as a promising approach for improving access to contextual information about genes in scientific literature. / För att analysera de stora mängder data som produceras med hjälp av next-generation sequencing krävs det att forskare har tillgång till och kan sammanställa information från tidigare forskning. I takt med att mängden data som finns tillgänglig i den vetenskapliga litteraturen ökar, så ökar även behovet av att använda AI och andra statistiska metoder för att få tillgång till denna data i analysen. I detta projekt utforskas kunskapsgrafer som verktyg för att tillgängliggöra kontextuell geninformation i vetenskapliga artiklar. Den explorativa metod som beskrivs i detta projekt är baserad på implementationen och jämförelsen av två olika tekniker för kunskapsgrafgenerering, en regelbaserad-statistisk metod samt en metod baserad på neurala-nätverk och co-occurrence, baserade på specifika kontexter inom litteraturen. Resultatet presenteras både i form av en kvantitativ jämförelse mellan metoder samt genom en kvalitativ expertutvärdering baserad på det kvantitativa resultatet. Den kvantitativa jämförelsen antydde att jämförelsen mellan kunskapsgrafer genererade med hjälp av olika metoder kan bidra med värdefull information för tolkningen och kontextualiseringen av viktiga gener. Resultatet visade även på begränsningar hos vissa metoder, till exempel gällande skalbarhet samt den mängd och typ av information som kan extraheras. Men även att metrics baserade på överlappning av hörn och kanter, samt metrics som tar hänsyn till den globala topologin i grafer kan vara användbara i jämförelsen av, samt för att representera skillnader mellan biologiska kunskapsgrafer. Resultatet från den kvalitativa expertutvärderingen visade att kunskapsgrafer baserade på geninformation extraherad från vetenskapliga artiklar kan vara värdefulla verktyg för att identifiera forskningsbias gällande gener, samt framhävde viktiga utmaningar gällande normalisering av biologiska entiteter inom området kunskapsgrafsutveckling. Baserat på dessa fynd framstår automatisk kunskapsgrafsgenerering som ett lovande tillvägagångssätt för att förbättra tillgängligheten av kontextuell geninformation i vetenskaplig litteratur.

Next-generation sequencing (NGS) ermöglicht das molekulare Profiling von Zellen mit beispiellos hohem Durchsatz. Allerdings ist der Fokus oftmals auf proteinkodierende Proteine beschränkt, wodurch die vollständige Diversität des Transkriptoms übersehen wird. Nicht-kodierende RNA-Moleküle variieren stark in ihrer Biogenese, Struktur und Funktion, wodurch ihre unverzerrte Inklusion in die Analyse erschwert wird. Diese Promotion fokussiert sich auf das Verständnis nicht-kodierender RNA und navigiert durch drei aufeinander aufbauende Säulen in der Analyse, um Beobachtungen in Wissen zu verwandeln: Generierung von Daten, Quantifizierung und Interpretation. Diese drei Säulen werden in den drei Kapiteln der Dissertation aus der bioinformatischen Perspektive adressiert, indem Schlüsselherausforderungen beschrieben und neue Lösungen vorgestellt werden, um die Analyse des gesamten Transkriptoms mit NGS-Techniken zu verbessern. Zunächst wird ein vollautomatischer Algorithmus vorgestellt, welcher die verschiedenen Quellen von aus der Vorberei- tung von Bibliotheken resultierenden Artefakten mittels unüberwachtes Lernen erkennt, was anschließend zur Optimierung der Protokolle zur Vorbereitung von total-RNA-seq-Bibliotheken genutzt werden kann. Zudem werden die primären Herausforderungen der Quantifizierung von total-RNA-seq behandelt: die Prozessierung von Reads, die mehreren, möglicherweise überlappenden Loci zugeordnet werden können, wie auch die Tatsache, dass manche Loci mehrfach im Genom vorkommen und ein Read zu all diesen Loci passen kann. Diese beiden Fälle können auch gleichzeitig vorkommen, was die Analyse von nicht-kodierender RNA mit üblichen Methoden erschwert. Um diese Problematik anzugehen, wird eine neue Software namens Multi-Graph count (MGcount) vorgestellt. Diese ordnet hierarchisch Reads Transkripten zu, um unter anderem eine Diskrepanz zwischen der Loci-Länge von small und long RNA zu berücksichtigen. Wenn Reads konsistent mehrfach alignieren, fasst MGcount Loci in Communitys zusammen. Es wird gezeigt, dass die Beurteilung der Expression auf der Community-Ebene eine genauere Quantifizierung von biologisch bedeutsamen RNA-Einheiten (Einfachtranskript oder Locusfamilien) ermöglicht. Schließlich wird MGcount angewandt, um nicht-kodierende RNA während der Differenzierung von induzierten pluripotenten Stammzellen in die Keimblätter Mesoderm, Endoderm und Ektoderm zu analysieren. In dieser Dissertation wird eine Multi-Omics-Analyse erfolgreich angewandt, um sowohl die Expressionsverläufe von verschiedenen RNA-Biotypen während der Determination zu charakterisieren als auch einen Zusammenhang bezüglich Chromatin-Remodellierung (“chromatin remodeling“) und DNA-Methylierung an den jeweiligen Loci herzustellen. Schlussendlich dient diese Dissertation als Ratgeber für alle Forschenden, die neue Einsichten in das nicht-kodierende Transkriptom gewinnen wollen. / Next-generation sequencing (NGS) techniques enable the molecular profiling of cells with unprecedented high throughput. Yet, in transcriptome analysis, the focus is often restricted to protein-coding RNA, overlooking the transcriptome in its entire diversity. Non-coding RNA molecules largely vary in biogenesis, structure and function and this challenges their unbiased inclusion into the analyses. This doctoral research places non-coding RNA understanding at the focus spot and navigates through the three workflow pillars that must align effectively to turn observations into knowledge: data generation, quantification, and interpretation. Throughout three chapters, this Thesis addresses these pillars from a Bioinformatics perspective, by outlining key challenges and introducing novel solutions to improve whole-transcriptome analysis through NGS techniques. First, we introduce a fully automatic algorithm that identifies sources of library preparation artifacts in an unsupervised manner and we demonstrate its utility within the development and optimization of total-RNA-seq library preparation protocols. Secondly, we address a major challenge in total-RNA-seq quantification; processing reads that align to multiple loci that overlap within the same genomic region or/and multiple loci that are present in high copy numbers. Such ambiguous alignments commonly arise due to the inherent characteristics of non-coding RNA. To tackle this, we introduce a novel software, named Multi-Graph count (MGcount), that hierarchically assigns reads to transcripts to account for loci length disparity between small-RNA and long-RNA and subsequently collapses loci where reads consistently multi-map into communities defined in a data-driven fashion. We show that these cohesive communities allow the quantification of biologically meaningful RNA entities (single-transcripts or locus-families) and estimate their abundance more accurately. Finally, we apply the developed method to investigate non-coding RNA in early development, specifically during the differentiation of Induced Pluripotent Stem Cells into the three germ-layer lineages, namely, mesoderm, endoderm, and ectoderm. In this study, we leverage a multi-omics analysis to characterize the expression trajectories of diverse RNA biotypes along cell-commitment and the interplay with chromatin remodeling and DNA methylation patterns at the locus surroundings. Ultimately, this work is intended to serve as a guide for all those who want to gain new insights from the non-coding transcriptome.

CoenzymeQ10 (CoQ10) functions as an electron carrier in mitochondria transporting electrons from CI and CII to CIII in the respiratory chain (RC) for normal cellular energy (ATP) production. Mutations in genes of a complicated and not yet well understood CoQ10 biosynthesis cause primary CoQ10 deficiency, a rare autosomal recessive mitochondrial disorder (MD) with diverse heterogeneous clinical phenotypes. Although the major function of CoQ10 is to serve as electron transfer molecule it furthermore possesses multiple metabolic functions which can result in secondary CoQ10 deficiency. Five main clinical phenotypes are associated with CoQ10 deficiency although distinct genotype-phenotype associations are still absent due to the limited molecular genetic diagnoses of most reported CoQ10 deficiency cases. A correlation was found between reduced levels of CoQ10 in muscle tissue and deficient CII + III RC enzyme activities in a South African patient cohort, the current indicators for potential CoQ10 deficiency. The aim of the study was therefore to identify nuclear-encoded mutations in genes associated with CoQ10 deficiencies in a cohort of South African patients diagnosed with respiratory chain deficiencies (RCDs). A high throughput target enrichment strategy was performed in order to identify previously reported and/or possible novel CoQ10-assosciated disease-causing variants using Ion Torrent next generation sequencing (NGS) and an in-house developed bioinformatics pipeline. The data obtained were compared to clinical presentations of the patients to interpret the results of the identified variants considered to be possibly pathogenic. Targeted genes associated with primary CoQ10- and secondary CoQ10 deficiency was successfully sequenced in 24 patients, identifying 16 possible disease-causing variants. Of these variants three compound heterozygous variants were identified in three patients in genes ETFDH, COQ6 and COQ7, which were considered to be pathogenic according to the available data provided. Further validation of these three variants supported its pathogenicity in at least two of these variants (ETFDH and COQ6). In conclusion: This study contributed to better understanding the aetiology of a South African cohort of patients diagnosed with MDs. It also highlighted the valuable role of NGS for such investigations, and furthermore identified areas in the biochemical and molecular diagnostic strategy where improvements could be made in the future. / MSc (Biochemistry), North-West University, Potchefstroom Campus, 2015