Theses and Dissertations from UMD
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New submissions to the thesis/dissertation collections are added automatically as they are received from the Graduate School. Currently, the Graduate School deposits all theses and dissertations from a given semester after the official graduation date. This means that there may be up to a 4 month delay in the appearance of a give thesis/dissertation in DRUM
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Item Quantifying the relative contribution and furthering qualitative understanding of ftz cis-regulatory elements in Drosophila melanogaster(2022) Fischer, Matthew Douglas; Pick, Leslie; Entomology; Digital Repository at the University of Maryland; University of Maryland (College Park, Md.)Embryonic development is coordinated by interactions within gene regulatory networks. This process is orchestrated at the level of transcription through the regulatory properties of enhancers, which direct spatiotemporal expression patterns when bound by specific trans-acting factors. Though enhancers can act upon promoters located at great distances irrespective of orientation, the contributions from these cis-regulatory elements (CREs) are limited by insulators and/or tethering elements that organize chromatin architecture. Much research has been conducted towards understanding the coordination of the segmentation genes that pattern the basic body plan of the fruit fly, Drosophila melanogaster, during embryogenesis. The pair-rule genes (PRGs) of this pathway, such as fushi tarazu (ftz), are expressed in seven alternating stripes across the embryo. These PRGs are required for the development of body segments, and the mis-regulation of a single transcriptional domain can result in the loss of a segment. Here, I have investigated the ftz CREs to more precisely determine their sufficiency to direct expression within ftz stripe domains and their necessity for doing so in the native context of the gene. To investigate the sufficiency, I have generated 36 standardized reporter transgenes from 18 CREs, tested in both forward and reverse orientations. All CREs examined have been inserted into the same XbaI site of the reporter plasmid, and the transgenes have been inserted into the same genomic region. Through in situ hybridization experiments, I have determined that the qualitative patterns conferred by every CRE is orientation-dependent, and I have identified two putative insulators and/or tethering elements, proposed to explain this observation. To investigate their necessity, I targeted four genomic regulatory regions for precise deletion using the CRISPR/Cas9 system to generate seven deletion mutants. Though deletions were expected to cause lethality, most of the mutants are homozygous viable and fertile; only a mutant simultaneously removing two seven-stripe CREs was homozygous lethal. Quantitative gene expression analysis by fluorescent in situ hybridization chain reaction revealed that there is a critical threshold of ftz abundance required in each stripe for segmentation to proceed. In conclusion, I have determined that the ftz CREs are redundant and function together in a non-additive manner.Item Rachmaninoff's Performance Style As Heard In His Recordings: An Artistic And Technical Survey(2022) HENRY, DAVID; Gowen, Bradford; Music; Digital Repository at the University of Maryland; University of Maryland (College Park, Md.)This dissertation explores the piano recordings made by Sergei Vasilyevich Rachmaninoff, in particular the works he recorded multiple times, identifying the larger developmental arc of his playing during the twenty-four years of his latter career. We show that the common axes of technique and interpretation provide an insufficient lens through which to understand his changing style; instead, we propose an additional dimension of identity which better captures Rachmaninoff’s evolution as a performer. Furthermore, this point of view allows a separation of his recordings into three distinct periods. Rachmaninoff stands as a unique figure in modern musical history, reflecting an improbable convergence of events: 1. he was a world-renowned composer for the first half of his career. 2. upon the emergency of the Russian revolution in 1917, he emigrated with his family to America at the age of forty-five, industriously working to become one of the leading concert artists of his generation. 3. he was one of the first pianists to leave a recorded legacy. Appropriate historical and biographical context is provided to better understand Rachmaninoff and his times, and to place his recordings in a fuller context. We also present the current state of knowledge and availability of Rachmaninoff’s extant recordings, the various technologies by which they were made, and the recording companies with which Rachmaninoff chose to work. We describe the importance of matrix numbers, alternate takes, potentially unreleased recordings, piano rolls, and unintentional recordings. Rachmaninoff’s recordings currently exist across many fragmented sources, some difficult to learn of and acquire. This dissertation thereby provides a modern guide to these recordings in hope of aiding future scholars.Item Computational approaches for improving the accuracy and efficiency of RNA-seq analysis(2020) Sarkar, Hirak N/A; Patro, Robert; Computer Science; Digital Repository at the University of Maryland; University of Maryland (College Park, Md.)The past decade has seen tremendous growth in the area of high throughput sequencing technology, which simultaneously improved the biological resolution and subsequent processing of publicly-available sequencing datasets. This enormous amount of data also calls for better algorithms to process, extract and filter useful knowledge from the data. In this thesis, I concentrate on the challenges and solutions related to the processing of bulk RNA-seq data. An RNA-seq dataset consists of raw nucleotide sequences, drawn from the expressed mixture of transcripts in one or more samples. One of the most common uses of RNA-seq is obtaining transcript or gene level abundance information from the raw nucleotide read sequences and then using these abundances for downstream analyses such as differential expression. A typical computational pipeline for such processing broadly involves two steps: assigning reads to the reference sequence through alignment or mapping algorithms, and subsequently quantifying such assignments to obtain the expression of the reference transcripts or genes. In practice, this two-step process poses multitudes of challenges, starting from the presence of noise and experimental artifacts in the raw sequences to the disambiguation of multi-mapped read sequences. In this thesis, I have described these problems and demonstrated efficient state-of-the-art solutions to a number of them. The current thesis will explore multiple uses for an alternate representation of an RNA-seq experiment encoded in equivalence classes and their associated counts. In this representation, instead of treating a read fragment individually, multiple fragments are simultaneously assigned to a set of transcripts depending on the underlying characteristics of the read-to-transcript mapping. I used the equivalence classes for a number of applications in both single-cell and bulk RNA-seq technologies. By employing equivalence classes at cellular resolution, I have developed a droplet-based single-cell RNA-seq sequence simulator capable of generating tagged end short read sequences resembling the properties of real datasets. In bulk RNA-seq, I have utilized equivalence classes to applications ranging from data-driven compression methodologies to clustering de-novo transcriptome assemblies. Specifically, I introduce a new data-driven approach for grouping together transcripts in an experiment based on their inferential uncertainty. Transcripts that share large numbers of ambiguously-mapping fragments with other transcripts, in complex patterns, often cannot have their abundances confidently estimated. Yet, the total transcriptional output of that group of transcripts will have greatly-reduced inferential uncertainty, thus allowing more robust and confident downstream analysis. This approach, implemented in the tool terminus, groups together transcripts in a data-driven manner. It leverages the equivalence class factorization to quickly identify transcripts that share reads and posterior samples to measure the confidence of the point estimates. As a result, terminus allows transcript-level analysis where it can be confidently supported, and derives transcriptional groups where the inferential uncertainty is too high to support a transcript-level result.Item DISSECTING THE GENE REGULATORY FUNCTION OF THE MYC ONCOGENE WITH SINGLE-MOLECULE IMAGING(2020) Patange, Simona; Larson, Daniel R; Girvan, Michelle; Biophysics (BIPH); Digital Repository at the University of Maryland; University of Maryland (College Park, Md.)The MYC oncogene contributes to an estimated 100,000 cancer-related deaths annually in the United States and is associated with aggressive tumor progression and poor clinical outcome. MYC is a nuclear transcription factor that regulates a myriad of cellular activities and has direct interactions with hundreds of proteins, which has made a unified understanding of its function historically difficult. In recent years, several groups have put forth a new hypothesis that questions the prevailing view of MYC as a gene-specific transcription factor and instead envision it as a global amplifier of gene expression. Instead of being an on/off switch for transcription, MYC is proposed to act as a `volume knob' to amplify and sustain the active gene expression program in a cell. The scope of the amplifier model remains controversial in part because studies of MYC largely consist of cell population-based measurements obtained at fixed timepoints, which makes distinguishing direct from indirect consequences on gene expression difficult. A high-temporal, high-spatial precision viewpoint of how MYC acts in single living cells does not exist. To evaluate the competing hypotheses of MYC function, we developed a single-cell assay for precisely controlling MYC and interrogating the effects on transcription in living cells. We engineered `Pi-MYC,' an optogenetic variant of MYC that is biologically active, can be visualized under the microscope, and can be controlled with light. We combined Pi-MYC with single-molecule imaging methods to obtain the first real-time observations of how MYC affects RNA production and transcription factor mobility in single cells. We show that MYC increases the duration of active periods of genes population-wide, and globally affects the binding dynamics of core transcription factors involved in RNA Polymerase II transcription complex assembly and productive elongation. These findings provide living, single-cell evidence of MYC as a global amplifier of gene expression, and suggests the mechanism is by stabilizing the active period of a gene through interactions with core transcription machinery.Item Preserving the Narrative of 20th Century Art Song: A Guide for Instrumental Transcriptions of Vocal Music(2020) Spence, Luke Jonathan; Gekker, Paul C; Music; Digital Repository at the University of Maryland; University of Maryland (College Park, Md.)Art song, by nature, is a combination of media: text and music. Composers map these two domains onto each other using established correspondences known as conceptual metaphors. The outcome, known as a conceptual blend, manifests a different cognitive perception than if the domains are experienced separately. The purpose of this dissertation is to keep these blends intact when performing instrumental transcriptions of vocal music. Without the ability to utter words, the performer must consider how they can bring life to the narratives through their instrument. A comprehensive set of performance practice guidelines are discussed, largely for practical application to the trumpet. Accompanying this dissertation are seventeen video recorded transcriptions of 20th century art song from German, French, and American composers, which implement a solution to be used along with these guidelines: subtitle translations. The addition of subtitles not only allows for the audience to experience the narratives in real-time with the music, but also creates an opportunity for instrumentalists to perform vocal repertoire that has yet to be explored in great depth. The expressive atonal and twelve-tone works of the Second Viennese School, experimental and Transcendental works of Charles Ives, deep Symbolist poetry set by Lili Boulanger, and evocative, text-driven works of Libby Larsen can be brought to life in compelling ways with their narratives intact.Item THE CORNO D'AMORE - A BAROQUE TRANSCRIPTION PROJECT(2019) Drew, Justin Thomas; Miller, Gregory E; Music; Digital Repository at the University of Maryland; University of Maryland (College Park, Md.)Throughout my musical career, I have always enjoyed Baroque music, especially in a church setting. I have had some opportunities to experience this first hand on the horn, but realized that the Baroque repertoire available to a horn player is incredibly limited. Furthermore, there became a realization that French horn music students do not interact enough with Baroque music. While undergraduate Music Education and Performance majors seek a complete education and experience in performing all genres of music, horn players are often left with a void of music and techniques from the Baroque Period. It is also not common for students to own period instruments, nor is it common for university studios to own these instruments. The baroque horn also follows Baroque tuning A=415, which is far lower than the modern traditional tuning, which is A=440. Therefore, I have created four Baroque transcriptions for Horn and Organ to add to the horn repertoire. Two transcriptions were taken from Oboe concertos and two from Oboe D’amore concertos. All of these works were originally accompanied with small string ensemble and continuo. I chose Oboe and Oboe D’amore because the melodies were accessible for the modern day horn player, where flute, violin, viola, and cello features brushed up against virtuosity. These melodies also embody four different types of baroque style and melodic mastery. The project includes a professional recording, the sheet music of the transcriptions, and a CD cover with liner notes. The CD was recorded at Chevy Chase Presbyterian Church on November 10, 12, 17, and 25, 2019. The horn used was a Lukas Horn #4 with an Osmun Chicago cup and Geyer rim. The organ is a Rieger Tracker Organ designed by Josef von Glatter-Gotz in 1973-4, performed on by Julie Vidrick Evans. My engineer and producer was Neil Brown, through the recording company Arts Laureate. The arrangements were created using the music writing software, Sibelius. The cover photo is credited to Kyung Jung at Yellowhale Photography.Item BOLT, AN AP2/ERF TRANSCRIPTION FACTOR, REGULATES ABIOTIC STRESS AND DEFENSE RESPONSES IN ARABIODPSIS THALIANA(2016) Bouten, Roxane; Chang, Caren; Cell Biology & Molecular Genetics; Digital Repository at the University of Maryland; University of Maryland (College Park, Md.)Biotic and abiotic stresses negatively affect plant growth and development, hence decrease productivity. Many AP2/ERF family transcription factors in plants have important roles in stress response signaling although most have not yet been studied. Here I show that expression of an Arabidopsis thaliana AP2/ERF family member, which I call BOLT, is regulated by a MAPK pathway that includes MEKK1, MKK1, MKK2, and MPK4, and has roles in both biotic and abiotic stress response as well influencing growth and development. In this thesis, I examined BOLT’s gene expression pattern and protein localization, using GUS and YFP reporter genes respectively, measured its expression in response to biotic and abiotic stress and plant hormones using RT-qPCR, examined phenotypes by generating overexpressing and RNAi lines, and analyzed its effect on downstream gene expression using a microarray at time points after inducing BOLT expression. I found that BOLT is expressed in various plant tissues and the protein localizes to nuclear bodies as demonstrated in onion epidermal cells. Knockdown (RNAi) plants exhibit greater drought tolerance and are larger than wild type under low light conditions, while the overexpressors exhibit a dramatic early flowering phenotype and are small and weak under low light. Gene expression analysis suggests BOLT regulates genes involved in photosynthesis, hormone biosynthesis and signaling, and defense, many of which are also regulated in the MAPK pathway. Increased BOLT expression downregulates two discreet systems, cyclic electron flow and glycine cleavage, components of photosynthesis and photorespiration, respectively, which are two systems that are important under low light conditions. Taking these results together, I conclude that BOLT functions downstream of a stress responsive MAPK pathway and regulates a variety of growth- and stress-related genes necessary to balance growth and defense in response to biotic or abiotic stresses, or low light conditions.Item Transcribing Viola da Gamba Literature for the Modern Double Bass(2015) Alger, Shawn; Murdock, Katherine; Manzo, Anthony; Music; Digital Repository at the University of Maryland; University of Maryland (College Park, Md.)The double bass, in its modern incarnation, dates from the late nineteenth century, which means that any performer wishing to play solo music from an earlier era must resort to transcriptions. For modern bassists wishing to play music from the Baroque era, the options of existing transcriptions are severely limited. Currently a handful of composers dominate the landscape of baroque music for double bass, and the music tends to borrow from either violin or cello repertoire. The fact of the matter is that Baroque music is tremendously underrepresented when compared against the entire oeuvre of available music for the double bass. This dissertation will present a collection of transcriptions from viola da gamba literature in a variety of styles and genres in order to illustrate the potential this music has for expanding the baroque repertoire for double bass. The scope of this paper will include solo music with accompaniment, unaccompanied transcriptions, and music for two and four basses. In transcribing these works I have kept as close to the original manuscripts and publications as possible with regards to bowing and notation. Deviations from the original have been clearly marked so that modern performers may decide for themselves how faithfully to reproduce what the composer wrote. It will also serve as a starting point toward reinventing this wonderful body of music that has heretofore been taken for granted.Item LEUNIG, LEUNIG HOMOLOG, AND SEUSS ARE TRANSCRIPTIONAL CO-REPRESSORS THAT REGULATE FLOWER DEVELOPMENT, MUCILAGE SECRETION, AND PATHOGEN RESISTANCE(2009) Bui, Minh; Higgins, William J; Biology; Digital Repository at the University of Maryland; University of Maryland (College Park, Md.)Transcriptional repression is an important regulatory mechanism for development. My thesis focuses on dissecting the function of Groucho (Gro)/Transducin-Like Enhancer of split (TLE) family of transcriptional repressors in plant development. My work characterizes two Arabidopsis thaliana genes, LEUNIG (LUG), first discovered to repress transcription of the floral homeotic gene AGAMOUS (AG), and LEUNIG_HOMOLOG (LUH), a gene with the highest sequence similarity to LUG. To investigate the functional redundancy between LUG and LUH, I constructed and analyzed lug; luh double mutants, and concluded that both LUG and LUH repress AG expression in the flower, with LUG playing a more prominent role than LUH. The double mutant also revealed a previously unknown function of LUG and LUH in embryogenesis because lug-3; luh-1 double mutants are embryo lethal, while the single mutants develop normal embryos. During the course of this study, I developed a new genotyping method called Simple Allele-discriminating PCR (SAP), which is cost-effective, quick, and easy to perform. This method has greatly facilitated my research as well as others in the lab. A second part of my thesis addresses the role of LUG and LUH in other developmental processes besides flower development. My data indicate that these two genes, like their counter parts in fungi and animals, act as "global co-repressors" in various developmental and physiological processes. My thesis work revealed that both co-repressors, together with its interacting protein SEUSS (SEU), repress the Salicylic Acid (SA) pathogen defense pathway. Although lug-3, luh-1, and seu-1> mutants induced PR1 expression at higher levels than wild-type, only lug-3 and seu-1 mutants were pathogen resistant. Furthermore, LUH functions as a positive regulator in seed mucilage secretion, a process important for proper seed germination, hydration, and dispersal. I propose a possible connection between the defect in mucilage secretion and pathogen defense in luh-1 mutant plants and seeds, which places the foundation for further investigation and may uncover mucilage secretion as a major defense mechanism. My thesis has provided important insights into how transcriptional co-repressors regulate diverse developmental and physiological pathways in higher plants.Item Role of ubiquitination in Caenorhabditis elegans development and transcription regulation during spermatogenesis(2008-08-12) Kulkarni, Madhura D; Mount, Stephen; Smith, Harold; Cell Biology & Molecular Genetics; Digital Repository at the University of Maryland; University of Maryland (College Park, Md.)Regulation of gene function can be achieved through a variety of mechanisms. In this dissertation, I present the genetic and molecular characterization of two genes involved in two distinct mechanisms of control. Each gene was initially identified by its functional role in sperm development in the model organism Caenorhabditis elegans. The first gene, uba-1, is an essential enzyme involved in protein turnover through ubiquitin-mediated proteolysis. A temperature-sensitive allele, (uba-1)it129, was isolated in a classical genetic screen for mutations that cause sperm-specific sterility. The second gene, spe-44, encodes a putative transcription factor. Its identification by microarray screening for sperm-enriched genes led to the cytological analysis of the deletion allele spe-44(ok1400), by reverse genetics approach. it129 encodes a conditional allele of uba-1, the sole E1 ubiquitin-activating enzyme in C. elegans. E1 functions at the apex of the ubiquitin-mediated conjugation pathway, and its activity is necessary for all subsequent steps in the reaction. Ubiquitin is covalently conjugated to various target proteins. Poly-ubiquitination typically results in target protein degradation, which provides an essential mechanism for the dynamic control of protein levels. Homozygous mutants of uba-1(it129) manifest pleiotropic phenotypes, and include novel roles for ubiquitination in sperm fertility, control of body size, and sex-specific development. We propose a model whereby proteins normally targeted for proteasomal degradation instead persist in uba-1(it129ts) and impair critical cellular processes. The second gene, spe-44, was identified as a putative sperm gene regulator in C. elegans based on its up-regulated expression during spermatogenesis and its significant sequence homology to the DNA-binding SAND domain. Genetic analysis of a deletion allele of spe-44(1400) has revealed its functional role during sperm development. Cytological analysis of spe-44(ok1400) showed developmental arrest of spermatocytes prior to spermatid differentiation. spe-44 mRNA is expressed in a narrow spatial and temporal window, just prior to spermatocyte differentiation, consistent with its functional role during spermatogenesis. Future study will be directed to find putative targets of spe-44 and the mechanisms that regulate gene expression using microarray analysis and yeast-one hybrid screens. These studies will help to understand transcriptional regulatory aspects of spermatogenesis in C. elegans.