Thesis (M.Eng. and S.B.)--Massachusetts Institute of Technology, Dept. of Electrical Engineering and Computer Science, 2002. / Includes bibliographical references (p. 83-86). / DNA sequencing is the process of determining the sequence of chemical bases in a particular DNA molecule-nature's blueprint of how life works. The advancement of biological science in has created a vast demand for sequencing methods, which needs to be addressed by automated equipment. This thesis tries to address one part of that process, known as base calling: it is the conversion of the electrical signal-the electropherogram--collected by the sequencing equipment to a sequence of letters drawn from ( A,TC,G ) that corresponds to the sequence in the molecule sequenced. This work formulates the problem as a pattern recognition problem, and observes its striking resemblance to the speech recognition problem. We, therefore, propose combining Hidden Markov Models and Artificial Neural Networks to solve it. In the formulation we derive an algorithm for training both models together. Furthermore, we devise a method to create very accurate training data, requiring minimal hand-labeling. We compare our method with the de facto standard, PHRED, and produce comparable results. Finally, we propose alternative HMM topologies that have the potential to significantly improve the performance of the method. / by Petros T. Boufounos. / M.Eng.and S.B.

by Sharon, Chui-Wah Luk. / Thesis (Ph.D.)--Chinese University of Hong Kong, 1997. / Includes bibliographical references (p. 128-146). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Mode of access: World Wide Web.

High-throughput DNA sequencing technology has advanced rapidly in the past few decades and is the driving force for personalized precision medicine. In this Thesis, a set of novel disulfide linker-based nucleotide reversible terminators (NRTs) has been designed and synthesized for application in DNA sequencing by synthesis (SBS), which is the dominant sequencing platform. The design and synthesis principles are outlined as follows. Four nucleotides (A, C, G, T) are modified as NRTs for the DNA extension reaction catalyzed by polymerase by attaching a cleavable fluorophore to a specific location on the base and blocking the 3′-OH group with a small chemically-reversible moiety so that the resulting molecules are still recognized by DNA polymerase as substrates. In these fluorescent NRTs, the fluorophores are attached through a disulfide (-SS-) cleavable linker to the 5-position of cytosine and thymine, and to the 7-position of deaza-adenine and deaza-guanine, and a small disulfide moiety is used to cap the 3'-OH group of the deoxyribose. The resulting fluorescent NRTs (3′-O-tert-butyldithiomethyl-dNTP-SS-fluorophores) are shown to be good substrates in DNA polymerase catalyzed reactions. The fluorophore and the 3′-O-tert-butyldithiomethyl group on a DNA extension product, which is generated by incorporating the 3′-O-tert-butyldithiomethyl-dNTP-SS-fluorophore in a polymerase reaction, are removed simultaneously and rapidly by treatment with a reducing agent, tris (3-hydroxypropyl) phosphine, in aqueous buffer solution. This one-step dual-cleavage reaction thus allows the reinitiation of the polymerase reaction and increases the SBS efficiency. DNA templates consisting of homopolymer regions were accurately sequenced by using this class of fluorescent nucleotide analogues on a DNA chip and a four-color fluorescent scanner. Compared with existing fluorescent NRTs, the unique disulfide linkers used to synthesize the NRTs described in this thesis are cleaved efficiently under DNA compatible conditions, leading to shorter scars on the DNA extension strand to further improvement of the DNA SBS technology.

This dissertation describes the design and synthesis of novel cleavable fluorescent/anchor modified nucleotide reversible terminators using 3’-O-dithiomethyl (3’-O-DTM; 3’-O-SS) as a linker to directly or indirectly attach a fluorescent reporter to achieve scarless DNA Sequencing by Synthesis (SBS). To develop these nucleotide analogues for four-color SBS, two nucleotide analogues (3’-O-ROX-SS-dATP and 3’-O-BodipyFL-SS-dTTP) with directly attached fluorescent dyes and two other nucleotide analogues with directly attached biotin or trans-cyclooctene (TCO) as anchors (3’-O-Biotin-SS-dCTP and 3’-O-TCO-SS-dGTP) were successfully designed and synthesized. The nucleotide analogues with a PEG-elongated linker (3’-O-ROX-PEG4-SS-dATP, 3’-O-BodipyFL-PEG4-SS-dTTP, 3’-O-Biotin-PEG4-SS-dCTP and 3’-O-TCO-PEG4-SS-dGTP) were also designed and synthesized to optimize their incorporation efficiency in polymerase reactions. In our design, Biotin and TCO were demonstrated to be anchor moieties with high efficiency and specificity for binding with fluorescently labeled streptavidin and tetrazine, respectively. The DNA extension products produced by polymerase incorporation of 3’-O-Biotin-SS-dCTP and 3’-O-Biotin-PEG4-SS-dCTP were accurately identified by binding to Cy5-labeled streptavidin, while the DNA extension products produced by polymerase incorporation of 3’-O-TCO-SS-dGTP and 3’-O-TCO-PEG4-SS-dGTP were identified with equal precision by reaction with TAMRA-labeled tetrazine. A proof-of-concept experiment was conducted to demonstrate four-color scarless SBS using the novel nucleotide analogues described above.

Title: Multiple sequence alignment using genetic algorithms Author: Zdeněk Pátek Department: Department of Software and Computer Science Education Supervisor: RNDr. František Mráz, CSc. Abstract: The thesis adresses the problem of multiple sequence alignment (MSA). It contains the specication of the proposed method MSAMS that allows to find motifs in biological sequences, to split sequences to blocks using the motifs, to solve MSA on the blocks and nally to assemble the global alignment from the aligned blocks and motifs. Motif search and MSA are both solved using genetic algorithms. The thesis describes the implementation of the method, conguration of its settings, benchmarking on the BAliBASE database and comparison to the ClustalW program. Experimental results showed that MSAMS can discover better alignments than ClustalW. Keywords: multiple sequence alignment, motif nding, genetic algorithms, ClustalW

The estuarine to fluvial sediments of the mid-Cretaceous (Late Albian/Early Cenomanian) Dakota Formation of Jefferson Co., Nebraska (NE) and Washington Co., Kansas (KS) were deposited in a marginal marine setting along the eastern margin of the Cretaceous Western Interior Seaway. Three depositional facies based on various lithic content are recognized in the study area: Facies 1: Fluvial Channel Facies, Facies 2: Paleosol/Interfluve Facies, and Facies 3: Bay Head Delta/Estuarine Facies. The facies interpretation helped confirm that the Dakota Formation was deposited in a marginal marine setting in which low-gradient fluvial systems supplied a wave-dominated, estuary system. Petrographic analysis of the Fluvial Channel Facies concluded that the sandstones can be classified as quartz-rich lithic arkose. These findings differ slightly from previous studies on Cenomanian Dakota Formation strata in Thurston Co., NE. Palynostratigraphic, subsurface, and sedimentologic evidence helped to delineate a more accurate sequence stratigraphic framework for the Dakota Formation in the study area. Three large-scale, unconformity-bounded, sequences (D0, D1, and D2) are recognized, within which deposits of the transgressive and falling stage systems tracts are preserved in the Dakota Formation in the study area. While no physical deposits exist for the falling stage and lowstand systems tracts, evidence for their past occurrence can be observed by the erosional nature of the sequence boundaries. Detailed analysis of the systems tracts framework allows delineation of a generalized sea-level curve for the Dakota Formation in the study area. Analysis of the sequence stratigraphic framework revealed a Late Albian/Early Cenomanian sea-level fall that subsequently created valley incisions of over 25 m into the Late Albian D1 sequence. A careful literature review combined with sequence stratigraphic evidence suggests that a geologically fast-acting eustatic sea-level mechanism lowered worldwide sea-levels by more than 25 m from Late Albian into Early Cenomanian time. A reevaluation of the mid-Cretaceous "greenhouse" world suggests that a glacioeustatic component to the observed sea-level changes may have occurred. A Southern Hemispheric polar ice sheet with limited extent and volume compared to "icehouse" continental ice sheets, along with global alpine glaciers fed by wet climate cycles are hypothesized to account for sea-level fluctuations that resulted in valley incision and subsequent filling in the study area.

Liminal sites are those on the verge of change, between boundaries and in a temporary state of ambiguity. Throughout my practice as an architect I was aware of the existence of such spaces. I was also aware that they were rarely the product of my intentional design effort. Because of that to me these spaces were precious. They represent moments in space of ambiguous function and questionable beauty but also moments I sought out everyday. This masters research is my way of refocusing my practice to engage with these types of spaces. The sense that this search will take me outside of my understanding of architecture lead me to chose to undertake it as a masters in landscape architecture. My main research question is: How can a designer construct a liminal site? The research concentrates on four central themes - development of a definition of the term

Thesis (Ph. D. in Chemical Engineering)--Vanderbilt University, Dec. 2007. / Title from title screen. Includes bibliographical references.

Thesis (Ph.D.)--University of Delaware, 2007. / Principal faculty advisor: Patrick M. Gaffney, School of Marine and Earth Studies. Includes bibliographical references.

Graduation date: 1995