Dissertations / Theses: 'SNP development'

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Colley, James P. M. "The development and application of SNP discovery technologies from 2005 to 2012." Thesis, Cardiff University, 2012. http://orca.cf.ac.uk/41990/.

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Single Nucleotide Polymorphisms (SNPs) are used as markers in association studies and may contribute directly to inherited disease. Here, we investigated high throughput in silico and in vitro methods for identifying SNPs and applied these to large scale genomic projects. We evaluated the utility of the Transgenomic NavigatorTM software to facilitate automated detection of aberrant denaturing high performance liquid chromatography (dHPLC) elution profiles. 3,747 dHPLC profiles were analysed with this software and 98.3% of products with profiles distinct from wild type harboured novel variants (Chapter 3). We applied this software to investigate whether rare inherited variants in genes that play a role in oxidative DNA damage repair (OxDR) predispose to colorectal adenomas (CRA) and found that unlike MUTYH, inherited variants in other OxDR genes are unlikely to be a frequent cause of CRA (Chapter 4). We evaluated the sequence analysis packages Sequencher, InSNP, Mutation Surveyor and Staden to identify variants in patients with CRAs (using >4,000 chromatograms). Staden and Mutation Surveyor correctly identified 76/77 (98.7%) SNPs and 96.7% and 99.3% of the genotypes respectively (Chapter 5). We compared an optimised version of Staden against Sequencher for variant detection over a 2.5kb region of the adenomatous polyposis coli (APC) gene in 969 healthy controls. We found 100% concordance between these packages and found that rare nonsynonymous variants in APC were significantly over-represented in patients with CRAs as compared to healthy controls (32/480 vs. 37/969, P=0.0166) (Chapter 5). We evaluated data held in dbSNP (build 129) for common variants in the Caucasian population by examining ten DNA repair genes and subsequently developed software for automatically extracting selected information from dbSNP (Chapter 6). This program was used to rapidly identify 221 common nonsynonymous SNPs in every DNA repair gene in the human genome; these were subsequently typed in 480 publically-available human lymphoblastoid cell lines generating a resource for functional analyses (Chapter 6). We also assessed the role of these SNPs in susceptibility to CRC and response to therapy by exploiting data and samples for the randomised controlled trial COIN (Chapter 7). Finally, we used Next Generation Sequencing (NGS) to discover SNPs in the exomes of 10 patients that exhibited peripheral neuropathy in response to chemotherapy and discovered that ERCC4 nonsense mutations may contribute to this toxicity (Chapter 8). NGS is likely to become the key SNP discovery technique over the next decade due to its potential to comprehensively search an entire genome at a comparatively low cost.

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Probst, Mario C. O. "Development and evaluation of multiplex and high-throughput SNP analysis for the ABCA1 gene." [S.l.] : [s.n.], 2004. http://deposit.ddb.de/cgi-bin/dokserv?idn=970904533.

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An, Chuanfu. "SNP CHARACTERIZAITON AND GENETIC AND MOLECULAR ANALYSIS OF MUTANTS AFFECTING FIBER DEVELOPMENT IN COTTON." MSSTATE, 2008. http://sun.library.msstate.edu/ETD-db/theses/available/etd-03302008-191842/.

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Cotton (Gossypium spp.) is the worlds leading textile fiber crop, and an important source of oil and protein. Insufficient candidate gene derived-markers suitable for genetic mapping and limited information on genes that control economically important traits are the major impediments to the genetic improvement of Upland cotton (G. hirsutum L.). The objectives of this study were to develop a SNP marker discovery strategy in tetraploid cotton species, SNP characterization and marker development from fiber initiation and elongation related genes, chromosomal assignment of these genes by SNP marker-based deletion analysis or linkage mapping, and genetic and molecular analysis of mutants affecting cotton fiber development. Phylogenetic grouping and comparision to At- and Dt-genome putative ancestral diploid species of allotetraploid cotton facilitated differentiation between genome specific polymorphisms (GSPs) and marker-suitable locus-specific polymorphisms (LSPs). By employing this strategry, a total of 222 and 108 SNPs were identified and the average frequency of SNP was 2.35% and 1.30% in six EXPANSIN A genes and six MYB genes, respectively. Both gene families showed independent and incongruent evolution in the two subgenomes and a faster evolution rate in Dt-genome than that in At-genome. SNPs were concordantly mapped to different chromsomes, which confirmed their value as candidate gene marker and indicated the reliability of SNP discovery stragey. QTL mapping by two F₂ populations developed from fiber mutants detected major QTL which explain 62.8-87.1% of the phenotypic variation for lint percentage or lint index in the vicinity of BNL3482-138 on chromosome 26. Single marker regression analyses indicated STV79-108, which was located to the long arm of chromosome 12 (the known location of N₁ and perhaps n₂ loci), also had significant association (R² % value 15.4-30.6) with lint percentage, lint index, embryo protein percentage and micronaire. Additional QTL and significant markers associated with other seed and fiber traits were detected on different chromosomes. Inheritance analysis indicated that both genetic models N₁N₁n₂n₂ and n₂n₂li₃lisub>3 could lead to the fiberless phenotype. The observation of fuzzless-short lint phenotype indicated fiber initiation and elongation were controlled by different mechanisms. The penetrance of Li₂ gene expression was observed in this study.

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Silva, Junior Orzenil Bonfim da. "Development and applications of high-throughput SNP genotyping technologies in non-model plant genomes." Universidade Cat??lica de Bras??lia, 2017. https://bdtd.ucb.br:8443/jspui/handle/tede/2273.

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In the last twenty-five years, we have witnessed the wide adoption of DNA markers for the study of genetic variation in many organisms. A DNA marker must have two or more identifiable allelic DNA sequences to be useful. It usually does not have a biological effect, but instead functions as a traceable landmark in the genome, found in a specific location, and transmitted by the standard laws of inheritance from one generation to the next. Its application goes beyond genetic mapping and includes the analysis of genetic diversity, marker-trait association studies, marker assisted selection and, more recently, with the advent of wholegenome sequencing, whole-genome association and genomic selection. Among the several types of DNA sequence polymorphisms that can be used as DNA marker, Single Nucleotide Polymorphisms (SNPs) are the most powerful for large-scale variation analysis. There are vast numbers of SNPs in every genome and they can be typed by methods that have been proven easy to automate. Detection of alternative alleles is rapid and effortless because it is based on well-known polymerase chain reaction and DNA oligomer hybridization assays. Various strategies have been devised to discriminate alleles at a SNP, including fixed DNA arrays technologies, solution hybridization techniques and many sequencing-based genotyping. In our study, we have developed high-throughput DNA marker systems for non-model, highly heterozygous, diploid tree species. We took advantage of the combined power of Next Generation Sequencing (NGS) technologies, well-established highly automated methods of SNP typing and bioinformatics algorithms to perform genome-wide DNA variation analysis. We used whole genome resequencing of pooled individuals to develop a high-density 60K SNP chip for Eucalyptus species (EucHIP60k) providing a 96% genome-wide coverage with 1 SNP/12???20 kbp, and 47,069 SNPs at ??? 10 kb from 30,444 of the 33,917 genes in the Eucalyptus genome. We then used high-density SNP data and whole-genome pooled resequencing to examine the landscape of population recombination (??) and theta (??), assess the extent of linkage disequilibrium (r2) and build the highest density linkage maps for Eucalyptus to date. Chromosome-wide ancestral recombination graphs allowed us to date the split of Eucalytpus grandis (1.7???4.8 million yr. ago) and identify a scenario for the recent demographic history of the species. In a final set of studies, we built the first genome assembly for a Neotropical forest tree, the Pink Ip?? (Handroanthus impetiginosus), a highly-valued keystone timber species. Genome sequence was screened for the development of a targeted-capture sequencing system for SNP genotyping consisting of nearly 24,000 probe sequences. This genotyping system showed flexibility as it allowed the identification of SNPs across different populations of the species in moderate sample sizes. The good genome coverage, consistent Ts/Tv ratio estimated across samples and fair technical reproducibility between replicates, in terms of recall and precision of the SNP calling and accuracy on genotypes, indicate that this genotyping platform can be confidently used to estimate population genetics parameters and carry out population genomics investigations at the genome-wide scale
***

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Halpern, Micah. "DEVELOPMENT AND FORENSIC APPLICATION OF DYE PROBE FLUORESCENCE RESONANCE ENERGY TRANSFER FOR IMPROVED DETECTION OF CHANGES IN DN." Master's thesis, University of Central Florida, 2008. http://digital.library.ucf.edu/cdm/ref/collection/ETD/id/2871.

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Discovering, screening, and associating changes in DNA sequence are important to a broad range of disciplines and play a central role in Forensic Science. The typical types of changes include sequence variations [single nucleotide polymorphisms (SNP)] and length variations [short tandem repeats (STR)]. The steps for forensic DNA sample processing are similar for both types of changes but diverge at the point of detection. A number of approaches are being explored for SNP genotyping while STR analysis primarily consists of size-based analysis by capillary electrophoresis. Limitations exist for all current detection methods that pose significant impacts to forensic analysis. Bi-allelic SNPs result in three possible genotypes with a minimal amount of information generated per marker. Limitations for SNP analysis are due to the inability to amplify a suitable number of SNP markers from low DNA content samples to provide an appropriate level of discrimination. Multi-allelic STR markers are currently the marker of choice for forensic typing but a variety of experimental artifacts are possible that consist of either biology or technology related causes. Molecular genotyping methods developed across other disciplines have potential to alleviate some of these shortcomings but no current approach is capable of genotyping both SNP and STR loci with a single chemistry. The need for a more effective, efficient, and generalized approach led to development of a unique method called Dye Probe Fluorescence Resonance Energy Transfer (dpFRET) and determination of its suitability for forensic analysis. The development phase of the research consisted of synthetic testing to establish proof of concept for the chemistry followed by polymerase chain reaction (PCR) based assays to demonstrate real world applications. Following successful development, the boundaries and limitations for the technology were established (sensitivity, allelic dropout, mixed samples) and efforts were made to improve the approach. In the process, parallel testing for other fields including molecular pathology and conservation biology were incorporated to explore potential widespread application of this new approach. The overall goal of this project was to develop and explore the limitations for a unique approach to genotyping both SNPs and STRs. A majority of the work involved development of the method itself with the ultimate objective of application for forensic science. The focus of this project was to address and alleviate some of the shortcomings of current approaches that result in potential limitations for forensic analysis. It is expected that future applications of this technology might impact a wide range of disciplines to aid in discovery, screening and association of changes in DNA sequence.
M.S.
Department of Chemistry
Sciences
Forensic Science MS

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Soliai, Marcus Makina. "De novo Genome Assembly and SNP Marker Development of Pyrenophora semeniperda." BYU ScholarsArchive, 2011. https://scholarsarchive.byu.edu/etd/2960.

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Pyrenophora semeniperda (anamorph Drechslera campulata) is a necrotrophic fungal seed pathogen of a variety of grass genra and species, including Bromus tectorum, an exotic grass that has invaded many natural ecosystems of the U.S. Intermountain West. As a natural seed pathogen of B. tectorum, P. semeniperda has potential as a biocontrol agent due to its effectiveness at killing dormant B. tectorum seeds; however, few genetic resources exist for this fungus. Here, the genome assembly of a P. semeniperda isolate using 454 GS-FLX genomic and paired-end pyrosequencing techniques is presented. The total assembly is 32.5 Mb and contains 11,453 gene models greater than 24 amino acids. The assembly contains a variety of predicted genes that are involved in pathogenic pathways typically found in necrotrophic fungi. In addition, 454 sequence reads were used to identify single nucleotide polymorphisms between two isolates of P. semeniperda. In total, 20 SNP markers were developed for the purposes of recombination assesment of 600 individual P. semeniperda isolates representing 36 populations from throughout the U.S. Intermountain West. Although 17 of the fungal populations were fixed at all SNP loci, linkage disequilibrium was determined in the remaining 18 populations. This research demonstrates the effectiveness of the 454 GS-FLX sequencing technology, for de novo assembly and marker development of filamentous fungal genomes. Many features of the assembly match those of other Pyrenophora genomes including P. tritici-repentis and P. teres f. teres, which lend validity to our assembly. These findings present a significant resource for examining and furthering our understanding of P. semeniperda biology.

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Rosén, Annie. "Development of pharmacogenetic tests and improvement of autosomal ancestry DNA test." Thesis, Linköpings universitet, Institutionen för klinisk och experimentell medicin, 2010. http://urn.kb.se/resolve?urn=urn:nbn:se:liu:diva-57977.

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This master thesis was performed at the personal genomics company DNA-Guide Europa AB. The goal was to create DNA tests for drug response and to update the already existing DNA test for autosomal ancestry. The DNA tests for drug response: The objective of this part of the master thesis was to create individual DNA test for response to each drug within different groups of medicines. The tests were meant to interest private customers. DNA-Guide uses a microarray technique for the DNA-analysis and this delimited the choice of SNPs. Inserts, deletions, repeats and copies of a whole gene can be difficult to implement on the microarray chip. The SNPs and studies used as a base for the tests had to fulfil several criteria. The studies must be large enough to prove that the association between the genotype and the response to the drug is valid among Europeans, since it’s the clientele of the company. The found association must also be strong enough to be of interest for a DNA test at DNA-Guide. If the SNPs could be implemented on the microarray chip a customer report was created about the possible results. The report had the same structure and design as those for the existing DNA tests at DNA-Guide. The work resulted in DNA tests and reports for medicines within the seven groups of medicines; anticoagulants, medicine against high cholesterol, blood pressure lowering medicine, asthma inhalers, antidepressants, birth-control pills and antiretroviral drugs. The DNA test for autosomal ancestry: The purpose of the update was to enhance to customers understanding of their results and the construction of the test. The update resulted in a description of how the used algorithm processes the results (from the DNA analysis) and a guide to interpret the results of the test. Conclusions: Both the DNA tests for drug response and the updated DNA test for autosomal ancestry can add value for the customers at DNA-Guide. The DNA tests for drug response can offer an explanation to why a medicine does not have an effect or reveal if the customer has higher risk of adverse effects. Even though recommendations for dosage or treatment could not be provided in almost all of the created DNA tests, being aware of the higher risk can be the first step to avoid adverse effects. The update of the DNA test report for autosomal ancestry resulted in a better description of the algorithm and limitations of the test, which can enhance the customers’ understanding of their results.

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Krampe, Matthew Stephen. "Development of Single Nucleotide Polymorphism (SNP) Markers for Rapid, Inexpensive, and Standardized Identification of Pallid (Scaphirhynchus albus) and Shovelnose (S. platorynchus) Sturgeon Larvae." OpenSIUC, 2011. https://opensiuc.lib.siu.edu/theses/675.

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The purpose of this project was to develop inexpensive, standardized, and high throughput Single Nucleotide Polymorphism (SNP) markers that discriminate between pallid (Scaphirhynchus albus) and shovelnose (S. platorynchus) sturgeon for use as a larval identification tool. A total of 67 polymorphic sites was identified in DNA sequences from three genes: Recombination Activating Gene-1, Beta Actin, and Beta-2-Microglobulin. Allele frequencies from the 10 most variable SNPs were characterized for both pallid and shovelnose sturgeon in three geographically separated populations throughout the range of the pallid sturgeon. To create a standardized method of genotyping SNPs for larval pallid and shovelnose sturgeon, 5' nuclease allelic discrimination (TaqMan) assays were designed for two unlinked SNPs that exhibited the greatest allele frequency differences between species. A power analysis compared these SNP loci and their diagnostic power for species discrimination compared to sixteen microsatellite loci currently used for species discrimination (Schrey et al. 2007) One SNP locus was the most powerful marker for species identification in the upper and middle Missouri River. This study provides practical genetic tools for species discrimination between pallid and shovelnose that will facilitate understanding addressing questions that were previously too costly, labor intensive or technically challenging to answer.

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Misyak, Sarah A. "Development of a SNP Assay for the Differentiation of Allelic Variations in the mdx Dystrophic Mouse Model." Thesis, Virginia Tech, 2008. http://hdl.handle.net/10919/32325.

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The purpose of this study was to develop a SNaPshotÂ® assay to simultaneously discriminate between the dystrophic and wild type (wt) alleles in mdx mice. The mdx mouse is an animal model for Duchenne muscular dystrophy (DMD), a severe and fatal muscle wasting disease. To evaluate possible treatments and to carry out genetic studies, it is essential to distinguish between mice that carry the mutant dystrophic or wt allele(s). The current Amplification-Resistant Mutation System (ARMS) assay used to genotype mdx mice is labor intensive and sometimes fails to yield typing results, which reduce its efficiency as a screening tool. An alternative assay based on single nucleotide polymorphism (SNP) extension technology (i.e., SNaPshotÂ®) would be advantageous because its specificity and capability to be automated would reduce the labor involved and increase the fidelity of each assay. A SNaPshotÂ® assay has been developed that provides a robust and potentially automatable assay that discriminates between the wt and dystrophic alleles. The assay has been optimized to use: an undiluted DNA in the PCR, a 0.1 ÂµM PCR primer concentration, a full PCR product for the SNP extension reaction, a 50ÂºC annealing temperature for the SNP extension in accordance with standard SNaPshotÂ® conditions, and a 0.4 ÂµM concentration of the SNP extension primer. The advantages of the resultant SNaPshotÂ® assay over the ARMS assay include higher fidelity, robustness, and more consistent performance within and among laboratories, and reduced risk of human error.
Master of Science

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Groß, Theresa Elisa [Verfasser]. "Development of novel SNP panels for the application of massively parallel sequencing to forensic genetics / Theresa Elisa Groß." Köln : Deutsche Zentralbibliothek für Medizin, 2017. http://d-nb.info/1144184878/34.

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Fitak, Robert Rodgers. "Conservation Genomics of the Endangered Mexican Wolf and De Novo SNP Marker Development in Pumas using Next-Generation Sequencing." Diss., The University of Arizona, 2014. http://hdl.handle.net/10150/316929.

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Traditionally, conservation genetics has examined neutral-marker (e.g microsatellite) surveys to inform the conservation and management of species. The field expanded together with the expansion of molecular biology, primarily enabled by polymerase chain reaction (PCR) and DNA sequencing technologies. Recently, advances in genomics and bioinformatics, notably next-generation sequencing (NGS), have demonstrated the ability to further enhance conservation genetic assessments. As a result, conservation genetics is rapidly transforming into a field of conservation genomics. Although complete genome sequencing and analysis is still beyond the reach of many conservation genetic projects, researchers can benefit by producing large amounts of genetic data quickly for their species of interest, or by exploiting existing genomic data for a closely related species. The research presented below serves as an example of these two different approaches. First, I review the current state of conservation genomics, utilizing examples when appropriate to illustrate different techniques and approaches. Next, I describe the development of a tool using NGS that is useful for the rapid genetic analysis of pumas (Puma concolor) called PumaPlex. This work details the methods involved and will be useful for anyone interested in working with a species where little genomic data is available. The last three chapters focus on using an existing genomic tool for the domestic dog to analyze admixture, quantify inbreeding, and identify potential adaptive variation in the endangered Mexican wolf (Canis lupus baileyi). The results demonstrated the Mexican wolf has no significant recent ancestry from domestic dogs, and that several loci may potentially be effective in increasing fitness in the reintroduced population.

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Yang, Hee Jeong. "Development of SCAR marker linked to a root-knot nematode resistant gene in peanut." Texas A&M University, 2004. http://hdl.handle.net/1969.1/1242.

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Root-knot disease caused by Meloidogyne spp. is the most important nematode disease of peanut. Even though many management strategies have been applied to control this disease on peanut, resistance is the most recommendable. Marker-assisted selection has been used as a useful tool for screening of resistant individuals in segregating populations. However, it requires many laborious steps. Thus, there is a need for PCR - based markers, which are more practical, rapid, and efficient. In this study, we tried to develop a SCAR marker linked to root-knot nematode resistance locus in peanut based on the RFLP marker R2430E. The entire sequence of R2430E was 2217 bp and contained one putative open reading frame (ORF) of 713 nucleotides. Thirteen primers including 5 forward and 8 reverse primers were synthesized to sequence the entireR2430E. Based on the results of BLAST searches, R2430E appeared to encode an AAA ATPase containing von Willebrand factor type A (VWA) domain from Magnetococcus sp. MC-1 (106 bits). To determine if there is a portion of the R2430E that hybridizes only to a band co-segregating with the resistance locus, we generated 4 probes spanning different parts of the gene. Southern analysis using these probes revealed identical banding patterns for each probe. Therefore, we concluded that there is very limited if any sequence polymorphism between different alleles detected by the R2430E probe. Additionally, this conclusion is supported by the experiment in which we tested 25 primer pairs derived from the R2430E using genomic DNA from both resistance and susceptible genotypes. In this experiment, all primer pairs amplified identical PCR fragments, suggesting again that there is little or no sequence divergence between putative alleles as differentiated by southern blotting. To identify possible single nucleotide polymorphisms (SNPs) between polymorphic R2430E RFLP bands, we cloned several fragments that span the entire R2430E transcribed sequence. Surprisingly, no SNPs were identified in the transcribed region of this gene. We propose that polymorphism detected by this RFLP marker is outside of the R2430E.

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Fontenas, Laura. "Analyse fonctionnelle des gènes ndrg4 et elmo1 dans le développement du système nerveux périphérique chez le poisson zèbre." Thesis, Université Paris-Saclay (ComUE), 2015. http://www.theses.fr/2015SACLS060.

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Les cellules gliales qui forment les segments de myéline du système nerveux périphérique (SNP) sont appelées cellules de Schwann. Elles assurent aux nerfs un soutien fonctionnel et permettent une conduction rapide et efficace de l'influx nerveux. Cette fonction requiert une communication réciproque entre les neurones et les cellules gliales qui les entourent. Une perturbation de cette interaction entraine le plus souvent une situation pathologique comme les neuropathies périphériques dont la plus connue est la maladie de Charcot-Marie-Tooth. Cependant, les mécanismes qui conduisent à ces pathologies sont encore peu connus et leur compréhension demande au préalable l'élucidation des mécanismes physiologiques qui contrôlent le développement du SNP. Ce travail a consisté en l'analyse de nouvelles fonctions des gènes ndrg4 et elmo1, dans le développement du système nerveux périphérique, chez le poisson zèbre. Nous avons montré que ndrg4 (n-myc downstream regulated gene) est un facteur neuronal qui régule le développement de la myéline périphérique en contrôlant le regroupement des canaux sodiques aux nœuds de Ranvier et l'expression de la mbp. Ndrg4 semble moduler l'échange vésiculaire entre les axones et les cellules de Schwann, en contrôlant l'expression de certaines protéines de relargage vésiculaire comme SNAP25, membre du complexe SNARE.Ce travail décrit par ailleurs une nouvelle fonction de elmo1 (engulfment and cell motility 1) dans le développement du SNP du poisson zèbre, où il favorise la survie des neurones dans lesquels il est exprimé. Nous avons montré qu'elmo1 protège les neurones de l'apoptose et que cette fonction est contrôlée par la voie nétrine1/unc5b en amont de Rac1. De ce fait, elmo1 est requis pour le développement du ganglion de la ligne latérale postérieure et des axones qui en émergent pour donner un système nerveux fonctionnel. Ainsi, ces travaux contribuent à une meilleure connaissance du développement du SNP et élucident pour la première fois une nouvelle voie de signalisation requise pour la survie des neurones dans le SNP
The glial cells that form myelin segments in the peripheral nervous system (PNS) are called Schwann cells (SCs). They provide functional support for nerves and allow a fast and efficient conduction of the action potentials. This requires a bilateral communication between axons and the associated glial cells. Disruption of this interaction often leads to peripheral neuropathies e.g. Charcot-Marie-Tooth disease. However, the mechanisms underlying these diseases remain poorly known and their understanding requires, at first, the elucidation of the physiological mechanisms responsible for PNS development. This work consists of a functional analysis of two genes, ndrg4 and elmo1, in the PNS development, using the zebrafish model. We showed that the neuronal factor ndrg4 (n-myc downstream regulated gene 4) regulates nodes of Ranvier organization and mbp expression along the Posterior Lateral Line nerve (PLLn). This is achieved, most likely, by the ability of ndrg4 to modulate vesicular exchange between axons and SCs. Indeed, the expression of some key proteins involved in vesicle docking and release such as SNAP25, a member of the SNARE complex, are significantly dependent on ndrg4.Moreover, this work describes a novel role for neuronal elmo1 (engulfment and cell motility 1) in PNS development by promoting neuronal survival within the PLL ganglion. We showed that elmo1 has protective role against apoptosis and that its function is controlled by the netrin1/unc5b signalling upstream of Rac1 and independently of macrophages role in apoptotic clearance. Therefore, elmo1 is required for the proper development of the PLL ganglion and the axonal development of the PLLn. Thus, this study further contributes to our understanding of PNS development and unravels a novel molecular pathway required for neuronal survival in the PNS

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Saha, Mandal Arnab. "Computational Analysis of the Evolution of Non-Coding Genomic Sequences." University of Toledo Health Science Campus / OhioLINK, 2013. http://rave.ohiolink.edu/etdc/view?acc_num=mco1372349811.

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Merrill, Keith R. "Usage and Development of Molecular Markers for Investigation of the Population and Ecological Genetics of Bromus tectorum L." BYU ScholarsArchive, 2011. https://scholarsarchive.byu.edu/etd/2955.

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This thesis includes two studies: The first examined patterns of neutral genetic diversity within Bromus tectorum L. across the IMW region, and uses patterns of microsatellite (SSR) genotype distribution to make inferences about the respective roles of adaptively significant genetic variation, adaptive phenotypic plasticity, and facultative outcrossing in the ongoing invasion and recent range expansion of B. tectorum. It has been previously demonstrated that, due to extremely low outcrossing rates, it is possible to characterize individual genotypes of this species using four SSR loci. We sampled 20 individuals from each of 96 B. tectorum populations (classified by region and habitat) from throughout the IMW and used these SSR markers to characterize each individual. We found 131 four-locus SSR genotypes; however, the 14 most common genotypes collectively accounted for 79.2% of the individuals sampled. Individuals with certain SSR genotypes sorted strongly into warm or salt desert habitats (stringent habitats) and flowered earlier than individuals with genotypes from more mesic habitats, providing evidence of adaptively significant genetic variation associated with these genotypes. Other SSR genotypes were found across a wide range of habitats though they tended to be less prevalent in stringent habitats, providing evidence that adaptive phenotypic plasticity may be important for the distribution of some common genotypes. We observed very few heterozygous individuals, consistent with the highly inbreeding reproductive strategy of B. tectorum. Because specialist genotypes dominating recently invaded areas within the IMW region contained unique alleles, they are not likely to have resulted from recombination, leading us to doubt the role of facultative outcrossing as a significant mechanism facilitating the current range expansion of B. tectorum in the IMW.Previous research investigating the population and ecological genetics of Bromus tectorum L. in the North American invaded range has relied on either allozyme or microsatellite (SSR) genetic analyses, both of which have proven to have shortcomings. In order to overcome the issues associated with these other marker types, in the second study of this thesis we developed single nucleotide polymorphism (SNP) markers for B. tectorum by 1) obtaining normalized cDNA, 2) sequencing normalized cDNA using 454 sequencing, 3) aligning resultant contigs and looking for SNPs, 4) designing assays for SNP validation and genotyping using KASPar, 5) converting working KASPar assays for use with the Fluidigm EP1 platform using the 96.96 Dynamic ArrayTM IFC. Sequencing resulted in 1258041 reads, which assembled into 65486 contigs (20782 large contigs exceeding 500 base pairs). Using selection criteria of at least 10x coverage and 30% of the minor allele, 3333 putative SNPs were identified. We developed KASP assays for 255 putative SNPs, which resulted in 101 working polymorphic assays. Ninety-six assays were then successfully converted for use with KASP on the Fluidigm EP1 genotyping platform using 96.96 dynamic arrays.

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Naidu, Alecia Geraldine. "The development of a single nucleotide polymorphism database for forensic identification of specified physical traits." Thesis, University of the Western Cape, 2009. http://etd.uwc.ac.za/index.php?module=etd&action=viewtitle&id=gen8Srv25Nme4_9261_1297760101.

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Many Single Nucleotide Polymorphisms (SNPs) found in coding or regulatory regions within the human genome lead to phenotypic differences that make prediction of physical appearance, based on genetic analysis, potentially useful in forensic investigations. Complex traits such as pigmentation can be predicted from the genome sequence, provided that genes with strong effects on the trait exist and are known. Phenotypic traits may also be associated with variations in gene expression due to the presence of SNPs in promoter regions. In this project, the identification of genes associated with these physical traits of potential forensic relevance have been collated from the literature using a text mining platform and hand curation. The SNPs associated with these genes have been acquired from public SNP repositories such as the International HapMap project, dbSNP and Ensembl. Characterization of different population groups based on the SNPs has been performed and the results and data stored in a MySQL database. This database contains SNP genotyping data with respect to physical phenotypic differences of forensic interest. The potential forensicrelevance of the SNP information contained in this database has been verified through in silico SNP analysis aimed at establishing possible relationships between SNP occurrence and phenotype. The software used for this analysis is MATCH&trade
.

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Price, Erin Peta. "Development of novel combinatorial methods for genotyping the common foodborne pathogen Campylobacter jejuni." Queensland University of Technology, 2007. http://eprints.qut.edu.au/16601/.

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Campylobacter jejuni is the commonest cause of bacterial foodborne gastroenteritis in industrialised countries. Despite its significance, it remains unclear how C. jejuni is disseminated in the environment, whether particular strains are more pathogenic than others, and by what routes this bacterium is transmitted to humans. One major factor hampering this knowledge is the lack of a standardised method for fingerprinting C. jejuni. Therefore, the overall aim of this project was to develop systematic and novel genotyping methods for C. jejuni. Chapter Three describes the use of single nucleotide polymorphisms (SNPs) derived from the multilocus sequence typing (MLST) database of C. jejuni and the closely related Campylobacter coli for genotyping these pathogens. The MLST database contains DNA sequence data for over 4000 strains, making it the largest comparative database available for these organisms. Using the in-house software package "Minimum SNPs", seven SNPs were identified from the C. jejuni/C. coli MLST database that gave a Simpson's Index of Diversity (D), or resolving power, of 0.98. An allele-specific real-time PCR method was developed and tested on 154 Australian C. jejuni and C. coli isolates. The major advantage of the seven SNPs over MLST is that they are cheaper, faster and simpler to interrogate than the sequence-based MLST method. When the SNP profiles were combined with sequencing of the rapidly evolving flaA short variable region (flaA SVR) locus, the genotype distributions were comparable to those obtained by MLST-flaA SVR. Recent technological advances have facilitated the characterisation of entire bacterial genomes using comparative genome hybridisation (CGH) microarrays. Chapter Four of this thesis explores the large volume of CGH data generated for C. jejuni and eight binary genes (genes present in some strains but absent in others) were identified that provided complete discrimination of 20 epidemiologically unrelated strains of C. jejuni. Real-time PCR assays were developed for the eight binary genes and tested on the Australian isolates. The results from this study showed that the SNP-binary assay provided a sufficient replacement for the more laborious MLST-flaA SVR sequencing method. The clustered regularly interspaced short palindromic repeat (CRISPR) region is comprised of tandem repeats, with one half of the repeat region highly conserved and the other half highly diverse in sequence. Recent advances in real-time PCR enabled the interrogation of these repeat regions in C. jejuni using high-resolution melt differentiation of PCR products. It was found that the CRISPR loci discriminated epidemiologically distinct isolates that were indistinguishable by the other typing methods (Chapter Five). Importantly, the combinatorial SNP-binary-CRISPR assay provided resolution comparable to the current 'gold standard' genotyping methodology, pulsed-field gel electrophoresis. Chapter Six describes a novel third module of "Minimum SNPs", 'Not-N', to identify genetic targets diagnostic for strain populations of interest from the remaining population. The applicability of Not-N was tested using bacterial and viral sequence databases. Due to the weakly clonal population structure of C. jejuni and C. coli, Not-N was inefficient at identifying small numbers of SNPs for the major MLST clonal complexes. In contrast, Not-N completely discriminated the 13 major subtypes of hepatitis C virus using 15 SNPs, and identified binary gene targets superior to those previously found for phylogenetic clades of C. jejuni, Yersinia enterocolitica and Clostridium difficile, demonstrating the utility of this additional module of "Minimum SNPs". Taken together, the presented work demonstrates the potentially far-reaching applications of novel and systematic genotyping assays to characterise bacterial pathogens with high accuracy and discriminatory power. This project has exploited known genetic diversity of C. jejuni to develop highly targeted assays that are akin to the resolution of the current 'gold standard' typing methods. By targeting differentially evolving genetic markers, an epidemiologically relevant, high-resolution fingerprint of the isolate in question can be determined at a fraction of the time, effort and cost of current genotyping procedures. The outcomes from this study will pave the way for improved diagnostics for many clinically significant pathogens as the concept of hierarchal combinatorial genotyping gains momentum amongst infectious disease specialists and public health-related agencies.

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Smith, Scott Matthew. "Application of Genome Reduction, Next Generation Sequencing, and KASPar Genotyping in Development, Characterization, and Linkage Mapping of Single Nucleotide Polymorphisms in the Grain Amaranths and Quinoa." BYU ScholarsArchive, 2013. https://scholarsarchive.byu.edu/etd/3548.

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The grain amaranths (Amaranthus sp.) and quinoa (Chenopodium quinoa Willd.) are important seed crops in South America. These crops have gained international attention in recent years for their nutritional quality and tolerance to abiotic stress. We report the identification and development of functional single nucleotide polymorphism (SNP) assays for both amaranth and quinoa. SNPs were identified using a genome reduction protocol and next generation sequencing. SNP assays are based on KASPar genotyping chemistry and were detected using the Fluidigm dynamic array platform. A diversity screen consisting of 41 amaranth accessions showed that the minor allele frequency (MAF) of the amaranth markers ranged from 0.05 to 0.5 with an average MAF of 0.27. A diversity screen of 113 quinoa accessions showed that the MAF of the quinoa markers ranged from 0.02 to 0.5 with an average MAF of 0.28. Linkage mapping in amaranth produced a linkage map consisting of 16 linkage groups, presumably corresponding to each of the 16 amaranth haploid chromosomes. This map spans 1288 cM with an average marker density of 3.1 cM per marker. Linkage mapping in quinoa resulted in a linkage map consisting of 29 linkage groups with 20 large linkage groups, spanning 1,404 cM with a marker density of 3.1 cM per SNP marker. The SNPs identified here represent important genomic tools needed for genetic dissection of agronomically important characteristics and advanced genetic analysis of agronomic traits in amaranth and quinoa. We also describe in detail the scalable and cost effective SNP genotyping method used in this research. This method is based on KBioscience's competitive allele specific PCR amplification of target sequences and endpoint fluorescence genotyping (KASPar) using a FRET capable plate reader or Fluidigm's dynamic array high throughput platform.

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Stephens, Alex J. "The development of rapid genotyping methods for methicillin-resistant Staphylococcus aureus." Queensland University of Technology, 2008. http://eprints.qut.edu.au/20172/.

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Methicillin-resistant Staphylococcus aureus (MRSA) is an important human pathogen that is endemic in hospitals all over the world. It has more recently emerged as a serious threat to the general public in the form of community-acquired MRSA. MRSA has been implicated in a wide variety of diseases, ranging from skin infections and food poisoning to more severe and potentially fatal conditions, including; endocarditis, septicaemia and necrotising pneumonia. Treatment of MRSA disease is complicated and can be unsuccessful due to the bacterium's remarkable ability to develop antibiotic resistance. The considerable economic and public health burden imposed by MRSA has fuelled attempts by researchers to understand the evolution of virulent and antibiotic resistant strains and thereby improve epidemiological management strategies. Central to MRSA transmission management strategies is the implementation of active surveillance programs, via which unique genetic fingerprints, or genotypes, of each strain can be identified. Despite numerous advances in MRSA genotyping methodology, there remains a need for a rapid, reproducible, cost-effective method that is capable of producing a high level of genotype discrimination, whilst being suitable for high throughput use. Consequently, the fundamental aim of this thesis was to develop a novel MRSA genotyping strategy incorporating these benefits. This thesis explored the possibility that the development of more efficient genotyping strategies could be achieved through careful identification, and then simple interrogation, of multiple, unlinked DNA loci that exhibit progressively increasing mutation rates. The baseline component of the MRSA genotyping strategy described in this thesis is the allele-specific real-time PCR interrogation of slowly evolving core single nucleotide polymorphisms (SNPs). The genotyping SNP set was identified previously from the Multi-locus sequence typing (MLST) sequence database using an in-house software package named Minimum SNPs. As discussed in Chapter Three, the genotyping utility of the SNP set was validated on 107 diverse Australian MRSA isolates, which were largely clustered into groups of related strains as defined by MLST. To increase the resolution of the SNP genotyping method, a selection of binary virulence genes and antimicrobial resistance plasmids were tested that were successful at sub typing the SNP groups. A comprehensive MRSA genotyping strategy requires characterisation of the clonal background as well as interrogation of the hypervariable Staphylococcal Cassette Chromosome mec (SCCmec) that carries the β-lactam resistance gene, mecA. SCCmec genotyping defines the MRSA lineages; however, current SCCmec genotyping methods have struggled to handle the increasing number of SCCmec elements resulting from a recent explosion of comparative genomic analyses. Chapter Four of this thesis collates the known SCCmec binary marker diversity and demonstrates the ability of Minimum SNPs to identify systematically a minimal set of binary markers capable of generating maximum genotyping resolution. A number of binary targets were identified that indeed permit high resolution genotyping of the SCCmec element. Furthermore, the SCCmec genotyping targets are amenable for combinatorial use with the MLST genotyping SNPs and therefore are suitable as the second component of the MRSA genotyping strategy. To increase genotyping resolution of the slowly evolving MLST SNPs and the SCCmec binary markers, the analysis of a hypervariable repeat region was required. Sequence analysis of the Staphylococcal protein A (spa) repeat region has been conducted frequently with great success. Chapter Five describes the characterisation of the tandem repeats in the spa gene using real-time PCR and high resolution melting (HRM) analysis. Since the melting rate and precise point of dissociation of double stranded DNA is dependent on the size and sequence of the PCR amplicon, the HRM method was used successfully to identify 20 of 22 spa sequence types, without the need for DNA sequencing. The accumulation of comparative genomic information has allowed the systematic identification of key MRSA genomic polymorphisms to genotype MRSA efficiently. If implemented in its entirety, the strategy described in this thesis would produce efficient and deep-rooted genotypes. For example, an unknown MRSA isolate would be positioned within the MLST defined population structure, categorised based on its SCCmec lineage, then subtyped based on the polymorphic spa repeat region. Overall, by combining the genotyping methods described here, an integrated and novel MRSA genotyping strategy results that is efficacious for both long and short term investigations. Furthermore, an additional benefit is that each component can be performed easily and cost-effectively on a standard real-time PCR platform.

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Chakrabortty, Sharmistha. "SNPs and Indels Analysis in Human Genome using Computer Simulation and Sequencing Data." University of Toledo Health Science Campus / OhioLINK, 2017. http://rave.ohiolink.edu/etdc/view?acc_num=mco1501726874739045.

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Guiteras, Capdevila Montserrat <1980&gt. "Development of Organocatalytic stereoselective SN1 type reactions." Doctoral thesis, Alma Mater Studiorum - Università di Bologna, 2012. http://amsdottorato.unibo.it/4370/.

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The proposal in my thesis has been the study of Stereoselective α-alkylation through SN1 type reaction. SN1 type reaction involves a stabilized and reactive carbocation intermediate By taking advantages of stability of particular carbocations, the use of carbocations in selective reactions has been important. In this work has been necessary to know the stability and reactivity of carbocations. And the work of Mayr group has helped to rationalize the behaviour and reactivity between the carbocations and nucleophiles by the use of Mayr’s scale of reactivity. The use of alcohols to performed the stable and reactive carbocations have been the key in my thesis. The direct nucleophilic substitution of alcohols has been a crucial scope in the field of organic synthesis, because offer a wide range of intermediates for the synthesis of natural products and pharmaceutics synthesis. In particular the catalytic nucleophilic direct substitution of alcohols represents a novel methodology for the preparation of a variety of derivatives, and water only as the sub-product in the reaction. The stereochemical control of the transformation C-H bond into stereogenic C-C bond adjacent to carbonyl functionalized has been studied for asymmetric catalysis. And the field of organocatalysis has introduced the use of small organic molecule as catalyst for stereoselective transformations. Merging these two concepts Organocatalysis and Mayr’s scale, my thesis has developed a new approach for the α-alkylation of aldehydes and ketones through SN1 type reaction.
La proposta della mia tesi è stato lo studio stereoselettiva di α-alchilazione attraverso la reazione di tipo SN1. SN1 reazione tipo comporta un stabilizzato e reattivo carbocatione intermedio Prendendo vantaggi della stabilità dei carbocationi particolari, l'uso di carbocationi in reazioni selettive è stato importante. In questo lavoro è stato necessario conoscere la stabilità e reattività dei carbocationi. E il lavoro di Mayr gruppo ha contribuito a razionalizzare il comportamento e reattività tra i carbocationi e nucleofili con l'uso della scala Mayr di reattività. L'uso di alcool ai eseguito i carbocationi stabili e reattive sono state la chiave nella mia tesi. La sostituzione nucleofila diretta di alcoli è stata portata cruciale nel campo della sintesi organica, poiché offrono un'ampia gamma di intermedi per la sintesi di prodotti naturali e di sintesi farmaceutica. In particolare il catalizzatore sostituzione nucleofila diretta di alcoli rappresenta una nuova metodologia per la preparazione di una varietà di derivati, e l'acqua come unico sottoprodotto nella reazione. Il controllo stereochimica del legame CH trasformazione in legame C-C stereogenico adiacente al carbonile funzionalizzata è stata studiata per la catalisi asimmetrica. E il campo dell'organocatalisi ha introdotto l'uso di piccola molecola organica come catalizzatore per trasformazioni stereoselettive. L'unione dell'organocatalisi questi due concetti e la scala Mayr, la mia tesi ha sviluppato un nuovo approccio per la α-alchilazione di aldeidi e chetoni, attraverso reazione di tipo SN1

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Rybička, Josef. "Zákaznický vývoj v systému SAP." Master's thesis, Vysoká škola ekonomická v Praze, 2013. http://www.nusl.cz/ntk/nusl-163921.

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This thesis deals with SAP system from technological point of view and focuses on SAP NetWeaver platform. The main goal of this work is to describe and demonstrate custom development in SAP system. Secondary goal is to provide an overview of SAP NetWeaver platform and its capabilities and its important value for companies. The development is shown by creating an application in ABAP language and by using MMSP method. This thesis is complete guide for beginning SAP developers and contains tutorials about programming in ABAP and using its development tools.

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Dušek, Martin. "Implementace protokolu SIP." Master's thesis, Vysoké učení technické v Brně. Fakulta elektrotechniky a komunikačních technologií, 2011. http://www.nusl.cz/ntk/nusl-218958.

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This Master’s thesis deals in detail with the SIP protocol – a method of communication between two entities, various types of transmitted messages and their content. Few SIP libraries are introduced and two of them are used for development of an application for audio/video conference-calls. Compilation of OPAL and PTlib libraries for Windows 7 Professional (64bit) is described, and problems resulting from lack of information provided by authors. New improved “how to build” is presented. At the end, paper focuses on several ways of development of mentioned application.

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Lin, Hanyang. "Role of SWI/SNF chromatin remodeling complex in melanoma development." Thesis, University of British Columbia, 2010. http://hdl.handle.net/2429/26228.

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Human cutaneous malignant melanoma is an aggressive form of skin cancer, for its ability to metastasize rapidly and its resistance to conventional radiotherapy and chemotherapy. The mammalian SWI/SNF complex mediates ATP-dependent chromatin remodeling processes that are critical for transcriptional regulation, control of cellular processes, and involvement in DNA repair. Therefore, aberrant expression of SWI/SNF chromatin remodeling complex is involved in cancer development. To investigate the role of SWI/SNF complex in the development of melanoma, we used tissue microarray technology and immunohistochemistry to evaluate the expression of SNF5, the common core subunit, and BRG1, the ATPase subunit, in melanocytic lesions at different stages, and we analyzed the correlation between SNF5 and BRG1 expression and clinicopathologic variables and patient survival. In addition, we also investigated the role of SNF5 in nucleotide excision repair (NER), a type of DNA repair mechanism that removes ultraviolet-induced DNA lesions. We found that reduced SNF5 expression is significantly associated with melanoma progression and a worse patient survival, and that SNF5 is an independent prognostic factor for human melanoma. Furthermore, we showed that downregulation of SNF5 protein level in melanoma cell lines enhanced chemoresistance of melanoma cells. This suggests that SNF5 plays an important role in melanomagenesis and may serve as a promising prognostic marker for melanoma. BRG1 expression, on the other hand, was found to be increased in primary and metastatic melanoma compared to dysplastic nevi. Knockdown of BRG1 in human melanoma cell lines reduced cell proliferation due to G1 phase arrest. This suggests that BRG1 might play a role in the initiation stage of melanomagenesis. As for the role of SWI/SNF complex in NER, our current observation demonstrated that in human keratinocytes, SNF5 is required for efficient removal of CPD and is required for UV-induced histone acetylation. In human melanoma cells, SNF5 does not seem to play a major role in NER, for it is not required for removal of CPD and UV-induced global chromatin relaxation. Taken together, these data implicate that SWI/SNF complex plays an essential role in melanoma development and may serve as a promising therapeutic target for melanoma.

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Staskowski, Maureen, Ann Tyler, A. Lynn Williams, and Jill Naturkas. "Effect of Intensive Professional Development on SLP Implementation of EBP." Digital Commons @ East Tennessee State University, 2013. https://dc.etsu.edu/etsu-works/2053.

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Lakay, Elthea Trevolee. "SIP-based content development for wireless mobile devices with delay constraints." Thesis, University of the Western Cape, 2006. http://etd.uwc.ac.za/index.php?module=etd&action=viewtitle&id=gen8Srv25Nme4_9048_1182233050.

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SIP is receiving much attention these days and it seems to be the most promising candidate as a signaling protocol for the current and future IP telephony services. Realizing this, there is the obvious need to provide a certain level of quality comparable to the traditional telephone service signalling system. Thus, we identified the major costs of SIP, which were found to be delay and security. This thesis discusses the costs of SIP, the solutions for the major costs, and the development of a low cost SIP application. The literature review of the components used to develop such a service is discussed, the networks in which the SIP is used are outlined, and some SIP applications and services previously designed are discussed. A simulation environment is then designed and implemented for the instant messaging service for wireless devices. This environment simulates the average delay in LAN and WLAN in different scenarios, to analyze in which scenario the system has the lowest costs and delay constraints.

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Mirza, Amin Ruhul. "Developments in supported aqueous-phase catalysis." Thesis, University of Bath, 1999. https://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.311179.

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Cingo, Siphelele Sanele. "Development of SNAP-tag-based fusion proteins targeting HIV-1 viral reservoirs." Master's thesis, Faculty of Health Sciences, 2021. http://hdl.handle.net/11427/32576.

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Background Globally, the HIV/AIDS epidemic has cost over 35 million lives and approximately a further 37 million people are currently infected with HIV. In South Africa alone, more than 7 million people are HIV positive. Since the initiation of combination antiretroviral therapy (cART), viral replication can be supressed below the limit of detection by conventional testing. There is, however, no approved therapy for the cure of HIV. This is because HIV establishes viral reservoirs in memory CD4+ T-cells, where replication is low or arrested, allowing prolonged survival. Since there is little or no replication, a therapeutic strategy which targets the viral production and replication becomes ineffective and upon cessation of antiretroviral therapy a dramatic viral relapse occurs. The eradication of HIV, therefore, requires the targeted killing of the reservoir cells, or latency reversal followed by the prevention of further infection using cART. Targeting of cell-surface antigens for therapeutic purposes is the basis of immunotherapy. FDA-approved monoclonal antibodies such as Trastuzumab have been used to treat breast cancer via the human epidermal growth factor 2 (HER2) receptor. Immunotoxins (ITs) composed of an antibody fragment fused to apoptosis-inducing protein toxins targeting cellsurface antigens have been used for therapy of refractory leukaemia. The anti-CD22 recombinant IT Moxetumomab pasudotox based on Pseudomonas aeruginosa exotoxin A (ETA) has been FDA approved to treat hairy cell leukaemia. Moxetumomab pasudotox targets the antigen CD22 found on the surface of tumour cells. The HIV neutralizing VHH-nanobody J3, isolated from an immunised Llama has demonstrated anti-HIV properties against more than 95 % of HIV strains in vitro. As part of an ongoing project to develop a J3-ETA IT, this work sought to produce a J3-SNAP fusion protein by osmotic stress expression in the presence of compatible solutes in the periplasmic space of E. coli. SNAP-tag is a self-labelling protein that covalently binds benzylguanine (BG)-modified substrates in a 1:1 stoichiometric ratio. When recombinantly fused to any protein of interest, SNAP-tag allows the stable labelling of the protein of interest of in vitro and in vivo imaging. The periplasmic space of bacteria has been reported as a dedicated compartment to express functional proteins of interest. Furthermore, osmotic stress expression in the presence of compatible solutes has been reported to result in up to a thousand-fold increase in protein yield for difficult to express proteins. This study ultimately aimed to understand whether a functional J3-SNAP or J3-ETA can be expressed under osmotic stress in the presence of compatible solutes, in the periplasmic space of E. coli. 11 Experimental work In this study, a SNAP-tag-based fusion protein and an ETA-based IT were designed using J3, an anti-HIV-1 Env VHH-nanobody isolated from an immunised llama. Using the SnapGene® software (v.5.0.8, GSL Biotech LLC, USA), in silico design and cloning of an ETA-based IT J3-ETA and SNAP-tag-based fusion protein J3-SNAP was performed. Molecular cloning of designed open reading frames (ORFs) was performed into appropriate bacterial expression plasmid vectors. Plasmid vectors confirmed to contain the required ORFs by Sanger sequencing were transformed into E. coli BL21-DE3. Histidine-tagged J3-SNAP was expressed by osmotic stress in the presence of compatible solutes. J3-SNAP was purified by IMAC and assessed by SDS-PAGE and Western blot analysis. To ascertain the binding of J3- SNAP to cells expressing HIV-1 Env in vitro, recombinant Env protein was transiently transfected into HEK293T-cells to generate an Env expressing cell line. Cell-surface binding of SNAP-Surface® Alexa Fluor® 488 -conjugated J3-SNAP on Env expressing HEK293Tcells was assessed by confocal microscopy analysis. Results Successful expression of J3-SNAP in E. coli BL21-DE3 was confirmed by SDS-PAGE and Western blot analysis. The J3-SNAP fusion protein was subsequently purified by IMAC. Purified J3-SNAP was conjugated to the benzyl guanine-modified fluorophore SNAPSurface® Alexa Fluor® 488 and full-length conjugated protein was confirmed by combinations of SDS-PAGE and Western blot analysis. Cell-surface binding of J3-SNAP to HIV-1 Env-expressing HEK293T-cells was demonstrated in vitro by confocal microscopy analysis. These results prompted the generation of the IT, J3-ETA, by replacing SNAP-tag with ETA. Conclusion Successful binding studies suggest using J3 to target HIV-1 Env. Accessing patient probes would allow for the confirmation of these results for future human applications. Future in vitro studies would need to confirm the selective elimination of Env expressing T-cells by J3-ETA and thereafter confirmed on Env-positive patient probes.

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Nguyen, Thinh. "CDX2 Regulates Gene Expression Through Recruitment of BRG1-Associated SWI/SNF Chromatin Remodeling Activity." Thesis, Université d'Ottawa / University of Ottawa, 2016. http://hdl.handle.net/10393/35377.

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The packaging of genomic DNA into nucleosomes creates a barrier to transcription which can be relieved through ATP-dependent chromatin remodeling via complexes such as the switch/sucrose non-fermentable (SWI/SNF) chromatin remodeling complex. The SWI/SNF complex remodels chromatin via conformational or positional changes of nucleosomes, thereby altering the access of transcriptional machinery to target genes. The SWI/SNF complex does not possess intrinsic DNA binding ability, and therefore its recruitment to target loci requires interaction with DNA-associated transcription factors. The Cdx family of homeodomain transcription factors (Cdx1, Cdx2 and Cdx4) are essential for a number of developmental programs in the mouse. Cdx1 and Cdx2 also regulate intestinal homeostasis throughout life. Although a number of Cdx target genes have been identified, the basis by which Cdx members impact their transcription is poorly understood. We have found that Cdx members interact with the SWI/SNF complex and make direct contact with Brg1, a catalytic member of SWI/SNF. Both Cdx2 and Brg1 co-occupy a number of Cdx target genes, and both factors are necessary for transcriptional regulation of such targets. Finally, Cdx2 and Brg1 occupancy occurs coincident with chromatin remodeling at certain of these loci. Taken together, our findings suggest that Cdx transcription factors regulate target gene expression, in part, through recruitment of Brg1-associated SWI/SNF chromatin remodeling activity.

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Langdon, Yvette Gabrielle Conlon Frank L. "A functional analysis of the role of SHP-2 in vertebrate heart development." Chapel Hill, N.C. : University of North Carolina at Chapel Hill, 2008. http://dc.lib.unc.edu/u?/etd,1551.

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Thesis (Ph. D.)--University of North Carolina at Chapel Hill, 2008.
Title from electronic title page (viewed Sep. 16, 2008). "... in partial fulfillment of the requirements for the degree of Doctor of Philosophy in the Department of Biology Molecular Cellular and Developmental Biology." Discipline: Biology; Department/School: Biology.

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Xu, Dan. "EFFECTS OF ACTIVATING MUTATIONS IN SHP-2 (PTPN11) PHOSPHATASE ON HEMATOPOIETIC CELL DEVELOPMENT." Case Western Reserve University School of Graduate Studies / OhioLINK, 2011. http://rave.ohiolink.edu/etdc/view?acc_num=case1295052361.

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DaBell, Alex McGregor. "Extent of Cysteine Modification of SNAP-25 In vitro." BYU ScholarsArchive, 2014. https://scholarsarchive.byu.edu/etd/4361.

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Exocytosis, the fusion of a vesicle to a cellular membrane, involves a protein named SNAP-25. This protein, containing two alpha helices connected with a linker region, is localized to the cell membrane via palmitic acids attached to the cysteine residues of its linker region in a process called palmitoylation. Are cysteine residues of the SNAP-25 linker region palmitoylated in an ordered manner and to a particular extent? The answer to this question may give insight into the regulated nature of exocytosis. While it is generally accepted that SNAP-25 must be palmitoylated in order to perform its exocytotic functions, the details surrounding this process are still being discovered, defined, and understood. In these studies we replicate the oxidation, reduction, and palmitoylation of SNAP-25 in vitro. Palmitoylating SNAP-25 in vitro, a process which occurs regularly in vivo, allows us to determine the extent of palmitoylation. In vitro palmitoylation of SNAP-25 was studied both with and without a native palmitoyl acyl transferase (PAT), DHHC-17, the enzyme to attach palmitic acids to cysteines in the linker region of SNAP-25. These studies were done under a variety of conditions designed to identify (1) components necessary for optimal palmitoylation and (2) extent of palmitoylation with components that mimic native conditions. Palmitoylation is a common modification for a variety of proteins, both soluble and membrane-bound. Like phosphorylation, palmitoylation is reversible and may play an important role in regulation of cellular processes. Specifically, the palmitoylation of SNAP-25 may play a critical role in the regulation of exocytosis and therefore learning further details about this important process may help us to better understand a variety of neurodegenerative diseases and states of decreased or compromised exocytosis.

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Birgersson, Erica. "Chemical methods to increase the reactivity of lignin : In the context of green chemistry and education for sustainable development." Thesis, KTH, Skolan för teknikvetenskaplig kommunikation och lärande (ECE), 2015. http://urn.kb.se/resolve?urn=urn:nbn:se:kth:diva-171044.

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The research concerning lignin in high value applications has increased during the last years due to its renewability and availability in the black liquor from pulp mills. Today the major part of kraft lignin found in the black liquor is used as fuel to gain energy in the recovery boiler. Lignin functions as natural glue in plants so that the function of kraft lignin as a phenol replacement in wood adhesives has been researched. Due to lignin's low reactivity the molecule must be modified prior to use. Demethylation is a method to increase the phenolic content in lignin to produce more reactive sites. Thiol mediated and iodine mediated demethylation was performed. Demethylated lignin was characterized by changes in phenolic and hydroxylic groups, molecular mass, elemental composition and other properties using methods including UV, SEC and 31 P NMR. The results showed a decrease in the phenolic content contrary to the increase that was expected. Really low yields were also gained which makes the results non-representative. Size evaluation showed that the percentage of high molecular content in the demethylated lignin sample had increased, which point towards the loss of low molecular mass fractions. Due to demethylation lignin may have been more hydrophilic and soluble in DMF and water. In addition to this bond cleavage may have produced smaller fragments which also increase the solubility. The results point towards the loss of smaller fragments in the DMF and water phases. The applied demethylation methods were evaluated in context of green chemistry. Production, waste, involving chemicals and efficiency were discussed and analyzed. The applied demethylation methods use DMF as solvent which is not a green alternative, greener solvents such as water or other energy adding methods could be used to make the process greener. The use ofNaOMe produces methanol as a byproduct which could be eliminated by using NaOH, future studies on the efficiency of the bases in the needs to be done. Nature science has a reputation of being hard and firm. By bringing in social issues in science education new ways of looking at science opens up. A social problem and at the same time an environmental problem in today's society is the large plastic mountain in the Pacific Ocean. An educational material of the "Samhallsfragor med Naturvetenskapligt Innehiill",SNI, (Societal issues with scientific content) principle has been evaluated with respect to the abilities that can developed together with whether students increase their science knowledge through this. The study showed that students can develop almost all abilities described in the curriculum and their knowledge in science by this type of material. Keywords Biomaterial, lignin reactivity, thiol mediated demethylation, iodine mediated demethylation, green chemistry, SNl-fall.
Forskning kring lignin i produkter har ökat under de senaste åren på grund av lignins fornyelsebarhet och tillganglighet i svartluten från massabruken. Idag används den största delen av sulfatligninet från svartluten som bränsle for att producera energi i sodapannan. Lignin fungerar som ett naturligt lim i växter och på grund av detta undersöks funktionen av kraftlignin som fenolersättning i trälim. Med anledning av ligninmolekylens låga reaktivitet behover lignin modifieras fore användning i produkter. Demetylering är en metod för att öka fenolhalten i lignin och skapa en högre reaktivitet. I denna studie utfördes Tiolmedierad och jodidmedierad demetylering. Det demetylerade ligninet utvärderades med avseende på forändringar i fenol-och hydroxylgrupper, molekylvikt, elementarsammansättning och andra egenskaper med hjälp av olika metoder,inklusive UV, SEC och 31 P NMR. Resultaten visade en minskning i fenolinnehall i motsats till den ökning som förväntades. Riktigt låga utbyten påvisades också vilket gör att resultaten inte är representativa. Storleksutvärdering visade att andelen med högt molekylviktsinnehåll i det demetylerade ligninproven hade ökat, vilket pekar mot förlust av lågmolekylära fraktioner. På grund av demetyleringen kan ligninet ha blivit mer hydrofilt och lösligt i DMF och vatten. Utöver detta kan bindningsklyvning ha skapat mindre fragment som också ökat lösligheten. Resultaten pekar mot förlust av mindre fragment i DMF-och vattenfaserna. De tillämpade demetyleringsmetoderna utvärderades med avseende på grön kemi. Produktion, avfall, kemikalier och effektivitet diskuterades och analyserades. De tillämpade demetyleringsmetoderna använder DMF som lösningsmedel, vilket inte är ett grönt alternativ. Grönare lösningsmedel såsom vatten, eller andra typer av energitillsättning, kan användas för att göra processen miljövänligare. Användandet av NaOMe i den thiolmedierade demethyleringen skapar metanol som en biprodukt vilket kan bytas ut mot vatten om NaOH istället används. Vidare studier behöver göras för att undersöka de båda baserna effektivitet. Naturvetenskapen har ett rykte om att vara hård och fast. Genom att föra in sociala frågor i den naturvetenskapliga utbildningen kan nya sätt att se på vetenskapen skapas. Ett samhälls problem och samtidigt ett miljöproblem i dagens sämhalle är det stora plast berget i Stillahavet. Ett undervisningsmaterial för"Samhällsfrågor Med Naturvetenskapligt Innehåll", SNI, principen har utvärderats med avseende på vilka förmågor som kan tränas tillsammans med huruvida eleverna kan öka sin vetenskapliga kunskap. Studien visade att eleverna kan utveckla nästan alia formågor som beskrivs i läroplanen och sina kunskaper inom naturvetenskapen genom denna typ av utbildningsmaterial. Nyckelord Biomaterial, ligninreaktivitet, tiol medierad demetylering, jodmedierad demetylering,grön kemi, SNI-fall.

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Ertl, Iris Elisabeth. "Functional interplay of two SWI/SNF chromatin-remodeling accessory subunits during C. elegans development." Doctoral thesis, Universitat Pompeu Fabra, 2015. http://hdl.handle.net/10803/286067.

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El complexos SWI/SNF són remodeladors de cromatina dependents d’ATP que es troben altament conservats al llarg de l’evolució. Aquests complexos poden regular l’accessibilitat a una zona genòmica alterant l’estat de la cromatina i actuant com a reguladors transcripcionals. A més d’una subunitat enzimàtica central i diverses proteïnes que conformen el nucli, els complexos SWI/SNF incorporen subunitats accessòries que confereixen especificitat i varien segons el tipus cellular i el context del desenvolupament. Les poteïnes accessòries humanes BAF60a, BAF60b i BAF60c representen proteïnes paràlogues amb funcions especialitzades, de manera que mutacions en elles s’han relacionat amb diverses malalties (p. ex. càncer). Per tal de tenir una millor visió de les funcions de les unitats accessòries del complex SWI/SNF hem estudiat els paràlegs de les proteïnes BAF, codificades pels gens ham-3 i swsn-2.2. Hem investigat les funcions de ham-3 i swsn-2.2 en diversos teixits i processos del desenvolupament i hem observat que els dos gens són redundants en diversos contextos. Tot i així, i com ocorre amb els gens humans, HAM-3 i SWSN-2.2 també han adquirit funcions específiques.
SWI/SNF complexes are ATP-dependent chromatin remodelers highly conserved through evolution. By altering the chromatin state, these complexes can regulate the accessibility of a given genomic region and thereby perform transcriptional regulation. Besides a central enzymatic subunit and various core proteins, SWI/SNF complexes incorporate accessory subunits that confer specificity to a given complex and vary depending on the cell type and developmental context. The human accessory proteins BAF60a, BAF60b and BAF60c represent paralog proteins with specialized functions, and mutations in the BAF60 genes are involved in human disease (e.g. cancer). To get closer insight in the functions of SWI/SNF accessory subunits, we studied C. elegans homologs of the BAF60 proteins, encoded by the paralog gene pair ham-3 and swsn-2.2. We investigated ham-3 and swsn-2.2 functions in various tissues and developmental processes and observed that the two genes act redundantly in many contexts. However, as their human counterparts, HAM-3 and SWSN-2.2 have also acquired specialized functions.

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Gellrich, Arne L. "Peace Plus the Shooting? : A Critical Evaluation of SDP Practices and Tenability." Thesis, Linnéuniversitetet, Institutionen för samhällsstudier (SS), 2014. http://urn.kb.se/resolve?urn=urn:nbn:se:lnu:diva-34533.

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The study discusses the phenomenon of Sport for Development and Peace, in short SDP, which in recent years and with active support from the United Nations has been constantly gaining importance. Focusing on football, as the most popular sport, the thesis asks the question whether the generally positive view on both sports and the effects of sport participation on behaviour and the psycho-social development of youths is indeed a realistic assessment and if, in consequence, the game of football is accordingly applicable to projects in a peace-building context. To answer that question, the thesis first gives an overview over existing views and agendas concerning SDP projects among the international community, NGO’s, the private sector and academia. Then, two case studies of projects in Israel and the Balkans are presented, followed by a review of academic findings on the overall impact of sports. In an analytical part, the findings on the views on SDP, the case studies and the research considering sport in general are brought together. As a main result, the study manages to answer the research question, reaching the conclusion that the ramifications of sport are indeed much more ambivalent than generally suggested, and that the assessment of NGO’s, MNC’s and the UN alike would need to be adjusted accordingly. The UN recommendation to further sports in a peace-building context is not supported, however the human right to access to sport is recognised and the proliferation of sports in this context encouraged. It is however pointed out that such programmes are not automatically conductive towards the aims of peace and development work, but should rather be closely watched and well planned and implemented to avoid negative effects. Furthermore, SDP projects should be more thoroughly connected to other civil society initiatives. Both recommendations are so far not followed by the initiatives selected for the case studies.

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Dyrebrant, Tobias. "Utveckling av användargränssnittet för Atlas Copcos portal för samarbete med underleverantörer (SCP)." Thesis, Örebro universitet, Institutionen för naturvetenskap och teknik, 2016. http://urn.kb.se/resolve?urn=urn:nbn:se:oru:diva-51592.

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Vid skapandet av ett användargränssnitt bör en utvecklare utgå från en rad designprinciper. Detta möjliggör ett effektivt användarflöde, där användaren navigerar genom systemet utan svårigheter. Därigenom kan användaren till fullo absorbera informationen som systemet erbjuder. Designprinciperna ligger till grund för det praktiska arbetet som utfördes för denna rapport. Syftet var att utveckla en webbportal som användes av ett företag för samarbete med dess underleverantörer. Genom att utgå från principerna, samt intervjuer med användarna, skulle förbättringar göras på det nuvarande systemet. Detta skulle resultera i en bättre användarupplevelse och en optimal effektivitet för systemet. Det praktiska arbetet delades upp i två steg. Det första steget innefattade konkreta ändringar på det nuvarande systemet, där mindre justeringar introducerades som enkelt kunde implementeras på den nuvarande webbportalen. Det andra steget handlade om en analys av marknadens främsta designlösningar, vilket skulle visa på existerande smarta och trendanpassade lösningar som kunde användas vid förbättring av systemet. Dessa steg utgjorde processen för det förbättringsarbete som skulle utföras på webbportalen. Resultatet blev ett användargränssnitt som tillfredsställer majoriteten av användare. Genom de objektiva designprinciperna skapades ett användarvänligt system med ett effektivt användarflöde.
When creating a user interface, a developer should base their work on a number of design principles. This enables an effective user flow, where the user navigates through the system without difficulties. Thereby the user can absorb the information that the system provides to the fullest. These design principles form the basis of the practical work that was carried out for this report. The purpose was to develop a web portal, which was used by a company for collaboration with its suppliers. By basing the development on the principles, as well as interviews with the users, improvements were to be made on the current system. This should result in a better user experience and an optimal efficiency of the system. The practical work was divided into two stages. The first stage involved concrete changes on the current system, where smaller adjustments were introduced that could easily be implemented on the current web portal. The second stage was about an analysis of the market's leading design solutions, which should show existing smart and up-to-date solutions that could be used to improve the system. These steps constituted the process of improvement work that was to be carried out on the web portal. The result was a user interface that satisfies the majority of users. Through the objective design principles a user friendly system with an efficient user flow was created.

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Balíčková, Alžběta. "Řízení výzkumných a vývojových projektů v PS modulu informačního systému SAP." Master's thesis, Vysoké učení technické v Brně. Fakulta podnikatelská, 2015. http://www.nusl.cz/ntk/nusl-224806.

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This master thesis focuses on the analysis and design of accounting, tax and legal methods of research and development projects. In the first part of this thesis are mention a theoretical background of research and development projects. Another part deals with the management of research and development projects. The following part is a financial and strategic analysis and analysis of the current status of research and development projects in the community. The design part is determined by the accounting, tax and legal methodology for managing research and development projects and is designed to control process research and development project under the previous methodology and using SAP. In conclusion, the benefits of this methodology defined for the selected company.

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Guidi, Cynthia J. "The Role of the SWI/SNF Component INI1 in Mammalian Development and Tumorigenesis: a Dissertation." eScholarship@UMMS, 2003. https://escholarship.umassmed.edu/gsbs_diss/69.

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In vivo DNA is compacted tightly, via its association with histones and non-histone proteins, into higher-order chromatin structure. In this state, the DNA is refractory to the cellular factors that require access to DNA. The repressive nature of chromatin is alleviated in part by the action enzymes that modify chromatin structure. There are two major groups of chromatin modifying enzymes: those that post-translationally modify histones by the addition of small chemical moieties and those that utilize the energy derived from ATP hydrolysis to physically disrupt chromatin structure. The SWI/SNF enzyme belongs to this latter group. The SWI/SNF complex was identified originally in yeast. Several of its subunits are required for the expression of a subset of inducible genes. The ATPase activity is provided by the SWI2/SNF2 protein. In mammals, there are two biochemically separable SWI/SNF complexes that contain either BRG1 or BRM, both homologs of yeast SWI2/SNF2. The yeast and mammalian SWI/SNF complexes are able to disrupt the Dnase I digestion pattern of in vitro assembled mononucleosomes and arrays, as well as facilitate the accessibility of restriction nucleases and transcription factors. The mechanism by which SWI/SNF functions has yet to be elucidated. SNF5 is a component of the yeast SWI/SNF complex. It is required for sucrose fermentation and mating type switching. The mammalian homolog of Snf5 is SNF5/INI1. SNF5/INI1 was identified simultaneously by two groups as a protein that shares homology with Snf5 and via a yeast two hybrid assay as a protein that interacts with HIV integrase (INtegrase Interactor). INI1 is a component of all mammalian SWI/SNF complexes purified to date. In humans, mutations and/or deletions in INI1 are associated with a variety of cancers, including malignant rhabdoid tumors, choroid plexus carcinomas, medullablastomas, primitive neuralectodermal tumors, and some cases of leukemia. Furthermore, constitutional mutations within INI1in individuals presenting with these tumors support the role of INI1 as a tumor suppressor. In this thesis, we show that Ini1 also functions as a tumor suppressor in mice. Approximately 20% of mice heterozygous for Ini1 present with tumors. Most of these tumors are undifferentiated or poorly differentiated sarcomas with variable rhabdoid features. All tumors examined to date show loss of heterozygosity at the Ini1 locus. We also show that Ini1 is essential for embryonic development. Mice homozygous-null for Ini1die between days 4 and 5.5 post-fertilization due to an inability to adhere to their substratum, form trophectoderm, and expand their inner cell mass. We further characterize the function of Ini1 in tumor suppression by generating mice heterozygous for both Ini1 and either Rb or p53. While heterozygosity at the Ini1 locus appears to have no effect on the rate of tumorigenesis in Rb-heterozygous mice, many of the tumors arising in compound heterozygous mice present with an altered morphology. This finding suggests that Ini1 may contribute to tumor progression due to loss of Rb. In contrast, mice compound heterozygous for Ini1 and p53 show a marked reduction in the rate of tumorigenesis compared to p53-heterozygous mice. Furthermore, the tumor spectrum is altered in these compound heterozygous mice. These findings suggest that Ini1 may function normally to repress p53 activity. Lastly, we show that expression of the Ini1 tumor suppressor itself is regulated tightly. Tissues and cells heterozygous for Ini1 express roughly equivalent levels of Ini1 protein and mRNA as their wild-type counterparts. We further show that this compensation is mediated by an increase in the rate of transcription from the wild-type Ini1 allele. Moreover, when exogenous Ini1 is introduced into Ini1-heterozygous cells, expression from the Ini1 promoter is reduced. These data indicate that a compensatory mechanism exists to ensure that the steady-state levels of Ini1 are constant. In summary, research detailed in this thesis has contributed to our understanding of the regulation of Ini1 as well as the role this protein plays in mammalian development and tumor suppression.

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Weng, Qing Yu. "Characterizing the Role of the SWI/SNF Chromatin Remodeling Complex in the Development of Melanoma." Thesis, Harvard University, 2016. http://nrs.harvard.edu/urn-3:HUL.InstRepos:27007727.

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The SWI/SNF complex is a multiunit chromatin remodeling complex required for normal cell development. Mutations within SWI/SNF are associated with rapid tumor formation and aggressive progression of melanoma and other cancers. SWI/SNF dysregulation can alter gene expression globally across multiple oncogenic pathways. By bypassing signaling molecules targeted by current drugs, aberrant SWI/SNF complexes create unique challenges for cancer therapy. Given these challenges, we aim to define the molecular alterations in melanoma upon loss of SWI/SNF and determine whether its dysregulation produces characteristic dependencies on alternative epigenetic pathways. Identification of these synthetically lethal vulnerabilities may uncover novel epigenetic mechanisms in oncogenesis. We generated stable melanoma cell lines with precise knockdown efficiencies of SWI/SNF subunits using short hairpin RNAs and single cell cloning. Using this pool of SWI/SNF mutant cells, we assayed 100 compounds targeting epigenetic regulators. We identified multiple independent dependencies in SWI/SNF-deficient tumor cells vulnerable to synthetic inhibition. These targets also provide opportunities for synergistic combination with other treatment modalities. In addition, we show crosstalk between the SWI/SNF complex and MITF pathway that alters cancer cell phenotype. These findings offer insight into the poorly understood mechanisms by which SWI/SNF dysregulation contributes to melanomagenesis and uncover targetable vulnerabilities in these tumors.

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Papademetriou, Suzanne Andrea. "Role of the SHP-1 tyrosine phosphatase in the regulation of oocyte growth and follicle development." Thesis, University of Ottawa (Canada), 2002. http://hdl.handle.net/10393/6300.

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The tyrosine kinase Kit is expressed in oocytes and is involved in primordial germ cell (PGC) proliferation and oocyte growth. Our goal was to determine the involvement of the SHP-1 phosphatase in regulating PGC proliferation and oocyte growth by examining SHP-1 deficient motheaten mice. Ovaries from wild-type and motheaten mice were observed at 10--13 days of age and after transplantation under the kidney capsule of SCID mice. Ovaries were analyzed by histological and western analyses. SHP-1 was expressed in all ovarian cell types throughout follicle development. At 10--13 days, motheaten animals had smaller ovaries, increased numbers of PGCs and decreased granulosa cell proliferation compared to controls. After transplantation, both groups had formed large antral follicles in similar proportions. Interestingly, motheaten oocytes achieved larger sizes than controls. These results suggest that SHP-1 may interact with Kit to regulate PGC proliferation and oocyte growth, however, the loss of SHP-1 does not impair granulosa cell proliferation and follicle development.

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41

Baromey, Neth. "Ecotourism as a tool for sustainable rural community development and natural resources management in the Tonle Sap Biosphere Reserve /." Kassel : Kassel Univ. Press, 2008. http://d-nb.info/991252586/04.

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42

Morra, Erica, and Lisa Zenker. "Chapter 1: In Search of Innate Leadership : Discovering, Evaluating and Understanding Innateness." Thesis, Linnéuniversitetet, Institutionen för organisation och entreprenörskap (OE), 2014. http://urn.kb.se/resolve?urn=urn:nbn:se:lnu:diva-34622.

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Every individual is born with different natural competencies that can be honed by both voluntary and involuntary environmental stimuli. The response our genotype decides to make, if any, towards those stimuli, determines how well our competencies develop. Each person’s coding and variations of genes will result in unique qualities in their phenotype, or physical structure. As a result, a person has various traits that are displayed through their behavior. DNA is genetically shown to express itself through traits by up to 75%. This leaves a sort of buffer of around 25%. This region is available for us to adapt to our environmental stimuli. Your innate qualities will not reach their full potential without stimulation from the environment, in a leadership case, with education and training and therefore it can be argued that environmental exposure is necessary to fully expose the potentials and capabilities of an individual, rather than instill a new skill or develop a talent that was not existent before. Innate leadership is not a permanent state, on the contrary, it is a continuously adaptive situation demanding contextual evolutionary changes or resignation from the subject occupying the role. When the needs and demands of a society or era outweigh the relevance of the innate leaders' traits and competencies, an evolution of leadership is needed to maintain a positive relationship between all parties involved. As a result, the innate leader will begin to lose their innateness in their role and unless they evolve and adapt (because the two actions are not the same) to new contextual needs, their tenure as leader will begin to be detrimental and counter-functional. What we want to put forward is a real, universal and constructive understanding of what makes a human happy, motivated and productive and how an innate person in context is a much better solution in the short and long run, for those around them when put to a task.

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Lenhard, Klaus G. "International Participation in AOS Standards Development." International Foundation for Telemetering, 1989. http://hdl.handle.net/10150/614724.

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International Telemetering Conference Proceedings / October 30-November 02, 1989 / Town & Country Hotel & Convention Center, San Diego, California
During the current decade, international cooperation in space projects has become more and more popular and this trend is increasing. Initially, this involved only single missions with agencies flying payloads on other agencies' spacecraft. Later, this trend continued with international ventures, involving different agencies. In the immediate future, even more challenging scenarios are foreseen. The best known example and prime driver for such sophisticated missions will be the Space Station Freedom and its participating partners' spacecraft. Some of the international missions (ESA missions) are described briefly in this paper, in order to set the scene for a better understanding of the complex needs for standards within advanced orbiting systems. These ventures call for efficient means for cooperation and interoperability. Part of these requirements can be met by following international standards for space communications and space data systems. The Consultative Committee for Space Data Systems (CCSDS) undertook the task of integrating the space data systems requirements and developing appropriate recommendations for data systems standards for these Advanced Orbiting Systems (AOS). All international partners in the Space Station Freedom Program participated in the definition, development, and review of the AOS recommendations. The need for better cooperation in space communications via data relay satellite prompted the formation of a three party international panel called the Space Network Interoperability Panel (SNIP). An important aspect is the need for verification and validation of the concept and of the detailed technical recommendations. For the immediate future, special compatibility campaigns, involving the international agencies are planned in order to ensure the smooth application and functioning of the AOS recommendations.

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Bojler, Görling Martin. "Energy system evaluation of thermo-chemical biofuel production : Process development by integration of power cycles and sustainable electricity." Doctoral thesis, KTH, Energiprocesser, 2012. http://urn.kb.se/resolve?urn=urn:nbn:se:kth:diva-105814.

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Fossil fuels dominate the world energy supply today and the transport sector is no exception. Renewable alternatives must therefore be introduced to replace fossil fuels and their emissions, without sacrificing our standard of living. There is a good potential for biofuels but process improvements are essential, to ensure efficient use of a limited amount of biomass and better compete with fossil alternatives. The general aim of this research is therefore to investigate how to improve efficiency in biofuel production by process development and co-generation of heat and electricity. The work has been divided into three parts; power cycles in biofuel production, methane production via pyrolysis and biofuels from renewable electricity. The studies of bio-based methanol plants showed that steam power generation has a key role in the large-scale biofuel production process. However, a large portion of the steam from the recovered reaction heat is needed in the fuel production process. One measure to increase steam power generation, evaluated in this thesis, is to lower the steam demand by humidification of the gasification agent. Pinch analysis indicated synergies from gas turbine integration and our studies concluded that the electrical efficiency for natural gas fired gas turbines amounts to 56-58%, in the same range as for large combined cycle plants. The use of the off-gas from the biofuel production is also a potential integration option but difficult for modern high-efficient gas turbines. Furthermore, gasification with oxygen and extensive syngas cleaning might be too energy-consuming for efficient power generation. Methane production via pyrolysis showed improved efficiency compared with the competing route via gasification. The total biomass to methane efficiency, including additional biomass to fulfil the power demand, was calculated to 73-74%. The process benefits from lower thermal losses and less reaction heat when syngas is avoided as an intermediate step and can handle high-alkali fuels such as annual crops. Several synergies were discovered when integrating conventional biofuel production with addition of hydrogen. Introducing hydrogen would also greatly increase the biofuel production potential for regions with limited biomass resources. It was also concluded that methane produced from electrolysis of water could be economically feasible if the product was priced in parity with petrol.

QC 20121127

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Johns, Anna. "Towards the development of novel Aspergillus fumigatus targeted antifungals with an in-depth analysis of Sfp-PPTase, PptA." Thesis, University of Manchester, 2016. https://www.research.manchester.ac.uk/portal/en/theses/towards-the-development-of-novel-aspergillus-fumigatus-targeted-antifungals-with-an-indepth-analysis-of-sfppptase-ppta(5f2e1052-01b3-4fb2-84b9-5871538351db).html.

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Humans are continuously confronted by the threat of fungal infection. Estimates put the number of individuals infected by superficial fungal disease at about 1.7 billion. Additionally there are a significant proportion of invasive infections which are difficult to treat and lead to an estimated 1.5 million deaths each year. Aspergillus fumigatus is an opportunistic, fungal pathogen which can lead to a variety of disease manifestations, generally termed aspergillosis and account for more than 200,000 life threatening infections annually with mortality rates of up to 95%. The occurrence of fungal disease has increased significantly due to the expansion of the immune deficient population, particularly those who receive immunosuppressive therapies. This combined with the problems surrounding current therapeutic options for fungal disease such as a rise in the incidence of antifungal resistance, adverse side effects in patients and drug-drug interactions has led to an urgent need to discover and develop an innovative class of antifungal agents. This thesis will address this requirement by validating a potential drug target, PptA, for combating A. fumigatus infection and explore methods to identify novel antifungal target identification. PptA is a Sfp-type 4′-Phosphopantetheinyl transferase required for transfer and covalent tethering of 4’-phosphopantetheine from coenzyme A to a conserved serine residue within a peptidyl carrier domain of a protein substrate. This project outlines the many critical roles PptA plays within A. fumigatus, such as; involvement in the production of many virulence factors as well as the biosynthesis of the essential amino acid, lysine. Furthermore, it is demonstrated that PptA is vital for secondary metabolite production in A. fumigatus, growth in iron limiting conditions and virulence. Finally, the design of a high-throughput screening assay capable of identifying inhibitors of PptA enzymatic activity further demonstrate the suitability of this target. It is the combined effect of all these characteristics that makes PptA such a suitable candidate for an antifungal target. Additionally two validation methods are included which can be used to identify novel drug targets. The first method utilises chemically induced haploinsufficiency profiling, a technique which has been proven successful for drug target identification and determining mode of action of drugs in S. cerevisiae and C. albicans. The project has shown this technology can be successfully used in A. fumigatus. Furthermore, a new technique for in vitro parallel fitness screening using next generation sequencing was validated with a library of phosphatase deletion mutants. Bioinformatic analysis against strains exhibiting severe fitness defects allowed the identification of three phosphatases that have the potential of becoming successful antifungal drug targets.

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Aumann, Craig Alvan. "Development, parameterization and numerical solution of an unsaturated flow model for water in the sapwood of a Douglas-fir tree /." Thesis, Connect to this title online; UW restricted, 2001. http://hdl.handle.net/1773/6374.

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Kaur, Tarminder. "Sporting lives and "development" agendas : a critical analysis of sport and "development" nexus in the context of farm workers of the Western Cape." University of the Western Cape, 2016. http://hdl.handle.net/11394/4935.

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Philosophiae Doctor - PhD
This thesis is about the sporting lives of people who work and/or live at the commercial grape and wine farms of the Western Cape. Collectively referred to as farm workers, they are identified by the Western Cape Provincial Government as a priority group in need of "development". Over the past 15 or so years, proclamations and practices of "sport for development and peace" (SDP) have emerged as globally recognised phenomena, where sport is promoted as a tool to achieve a broad range of "development" objectives, including the Millennium Development Goals. As a research topic, SDP scholars examine the practical and theoretical usefulness of sport as a tool for addressing a diverse set of social, health, political and economic issues through education, diplomacy, inclusion, and awareness programmes. Instead of attending to the questions of whether or how sport might serve "development" ends, this study offers a critical analysis of the nexus between sport and "development" (SDN) in the context of farm workers of the Western Cape. Informed by James Ferguson‘s analysis of "development" as an 'anti-politics machine' (1990), I adopt a deconstructionist approach that examines issues beyond the narrow confines of "development" problems and programmes. As he argues, "development" continues to serve as a-central organising concept‘ to discuss and assess desired change in social and economic realms, which is evident in the programmes of farm worker "development" and how these continue to retain a place in the policy and political discourses on agrarian transformation in post-apartheid South Africa. With an appreciation of the Western Cape‘s agrarian history and politics and how they shape present-day farm labour conditions, I have critically analysed the discourses and practices of farm worker "development" and SDP in the light of broader structural realities, everyday sporting lives and the "development" experiences of farm workers. The central organising question of this thesis is: how do "development" problems and the solutions sought for in SDP discourses and programmes correspond to the social, economic and political realities of their subjects? Drawing on my ethnographic fieldwork conducted at farmlands in and around Rawsonville, a small rural town, from April 2012 to May 2013, I illustrate different and seemingly disconnected frames and positions from which theories of SDP and farm workers‘ experiences of sport and "development" were observed. The analysis is organised around three contrasting frames of observation, namely: 1) historical and contemporary discourses and politics of farm worker "development" and SDP programmes and practices, 2) structural arrangements of competitive and physical infrastructure for official sport, and 3) everyday (official and unofficial) sporting practices and experiences of the rural working class people. With a particular attention to continuities and contradictions in historical and contemporary farm worker "development" discourses and selected SDP case studies, I demonstrate that while SDP agendas directed at farm workers may serve divergent and at times conflicting interests, farm workers' own agency, initiative and aspirations do not feature in SDP programmes and broader "development" discourses. The contrasts and counter-narratives presented in discussing these case studies and stories complicate and contest simplified notions commonly projected in global SDP discourses and locally specific "development" agendas. Beyond the confines of sporadic and temporary SDP projects, there was a vibrant and active world of formal and informal sport among the farm workers of Rawsonville. By focusing on the everyday sporting lives of athletes, coaches, managers, organisers and soccer clubs, I paint a picture that reveals the diversity and inconsistency of experiences and meanings of farm worker as an identity, a class position and an occupation. Interrogating how farm workers were embedded within the broader rural sport structures, I describe the complex set of factors that shaped their experiences of, access to, and participation in, sport. I argue that while sport was passionately pursued irrespective of direct or corollary "development" benefits, it was unofficial and under-the-radar sport networks and practices that served as vital spaces of autonomy, initiative and self-realization, even for those who may not otherwise have had such opportunities. And while the politically disengaged and enthusiastically embraced qualities of sport may continue to be among the reasons for its traction in "development" and peace agendas, these very same qualities allow sport to be usefully employed as an ethnographic method. Among the formative turns I took in conducting and presenting my research observations was to implicate myself and invite the reader into the confusing and complex process of learning and knowledge production. By way of conclusion, I argue for refocusing the gaze of research on studying sport as part of the broader scope of subaltern sociality.

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48

Magugu, Freddy-Junior Siybaulela. "Development of potential immunodiagnostic & therapeutic techniques using SNAP-fusion proteins as tools for the validation of Triple-negative Breast Cancer." Master's thesis, Faculty of Health Sciences, 2021. http://hdl.handle.net/11427/32795.

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Globally, breast cancer is the leading cause of death in the female population aged 45 and below with a breast cancer incidence reaching 18.1 million in the year 2018. Triple negative breast cancer (TNBC) is part of a group of cancers that lack the expression of Progesterone receptor (PR), Estrogen receptor (ER) and Human epidermal growth factor receptor 2 (HER2). TNBC is commonly associated with early stage metastasis with low survival rates as well as a high frequency of recurrence and proves to be problematic in both the young and elderly female populations. Conventional diagnostic methods for TNBCs include mammography, magnetic resonance imaging (MRI) and ultrasound while therapeutic methods include mastectomy and breast conserving surgery (coupled with radiation therapy). The lack of effective therapeutic options, poor prognostic value and high rates of metastasis, has made treatment of TNBC difficult. The major focus of this work was on the following tumour associated antigens (TAAs): CSPG4 (a transmembrane protein found in 50% of TNBC cases), EGFR (which is overexpressed in 13-76% of TNBCs), and MSLN (which is overexpressed in 67% of TNBCs) as potential targets for monospecific therapy. The evolution of antibody-based immunotherapy strategies has led to applications of single chain variable fragment (scFv) & single domain/nanobody (VHH) antibody formats for diagnostic and therapeutic purposes. In this work, these recombinant antibody fragments have been combined with SNAP-tag, a modified version of the human DNA repair enzyme O6-alkylguanine-DNA-alkyltransferase (AGT), which autocatalytically binds benzyl-guanine modified substrates such as fluorophores or small molecule toxins covalently in a 1:1 stoichiometry. In this study, the primary aim was the comparison of different antibody formats fused to SNAPtag and the potential of these biopharmaceuticals towards immunodiagnosis and therapy of TNBCs. First functionalities of two scFv SNAP fusion proteins and one VHH SNAP fusion protein previously not having been described are provided through binding analyses on receptor positive tumour cell lines. This was achieved by in-silico design and molecular cloning of genetically fused antiCSPG4(scFv), -MSLN(scFv), -MSLN(VHH), -EGFR(scFv) & -EGFR(VHH) to SNAP-tag. The final constructs were confirmed by Sanger sequencing and subsequently transfected into a mammalian vector system (HEK293T) for transient expression of the engineered fusion proteins. Full length protein purified from cell culture supernatant was analysed for diagnostic/therapeutic activities dependant on the substrate attached in the form of a fluorophore or small molecule toxin resulting in recombinant antibody-drug conjugates (ADCs). The study shows promise in providing new immunodiagnostic and therapeutic agents that are specific and less harmful than the current state of the art procedure

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49

Getao, Shi. "DEVELOPMENT OF FUNCTIONAL PROTEOMICS APPROACHES FOR STUDYING RETROGRADE TRANSPORT." Phd thesis, Ecole Normale Supérieure de Paris - ENS Paris, 2011. http://tel.archives-ouvertes.fr/tel-00635924.

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Le trafic rétrograde permet le transport des protéines de la membrane plasmique vers le réticulum endoplasmique, via l'appareil de Golgi, évité la dégradation des lysosomes. Des études antérieures ont montré que le transport rétrograde est crucial pour les fonctions cellulaires telles que la signalisation, l'homéostasie ionique, et l'établissement de gradients du morphogène Wnt. Par ailleurs, le transport rétrograde joue un rôle essentiel dans l'internalisation cellulaire des facteurs pathogènes comme les toxines protéiques et les protéines de virus. Toutefois, la liste actuelle des protéines cargos est limitée, et il est probable que de nombreuses fonctions cellulaires du transport rétrograde restent encore à découvrir. De toute évidence, un fort besoin existe pour une caractérisation plus poussée de cette voie de transport. Dans cette étude, quatre différentes approches protéomiques ont été développées visant à identifier les protéines membranaires prenant la route du transport rétrograde: SNAP-tag, sulfatation, la FKBP, et la streptavidine. Parmi ceux-ci, l'approche SNAP-tag s'est avéré être la stratégie la plus efficace pour identifier les candidats du fret de la voie rétrograde. Cette stratégie est basée sur la modification covalente du protéome de la membrane plasmique avec un benzylguanine (BG) dérivés. Seules les protéines membranaires BG-taggées qui sont ensuite transportés par voie rétrograde peuvent coupler covalentement à une protéine de fusion de SNAP-tag localisée dans la TGN. L'approche a été validée, étape par étape, en utilisant une protéine cargo rétrograde bien étudiée, toxine Shiga sous-unité B (STxB). Nous avons pu montrer que les STxB peuvent être capturés et identifiées par l'approche SNAP-tag. Par ailleurs, couplé à la LC-MS, les candidats des nouvelle protéines de la voie rétrograde ont été identifiés. Les trois autres approches ont guidé notre choix vers la stratégie la plus efficace en protéomique, en apportant des possibilités d'expérimentation et de défauts dans les domaines de la chimie organique et en biologie cellulaire. Généralement, ce travail de thèse a permis de développer une nouvelle stratégie protéomique qui pourraient être appliquée pour identifier les nouvelles candidats de la route rétrograde. Nous avons mis au point une approche dynamique en protéomique qui complète la protéomique traditionnelle. Par ailleurs, le concept d'utiliser des outils chimiques pour étudier le transport rétrograde peut également être appliqué à d'autres voies d'endocytose.

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50

Gazit, Salomé. "Regulation of vascular development and homeostasis by platelet-derived Sphingosine 1-Phosphate." Thesis, Sorbonne Paris Cité, 2015. http://www.theses.fr/2015PA05T035.

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Dissertations / Theses on the topic 'SNP development'

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