Dissertation submitted in partial compliance with the requirements for a Masters Degree in Technology: Homoeopathy, Durban University of Technology, 2008. / Introduction This study investigated the effectiveness of homoeopathic dilutions of abscisic acid (ABA), molybdenum and allopurinol on inhibiting or promoting the germination of barley seeds (Hordeum vulgare cv. Stirling, ex Caledon, Western Cape, South Africa, 1998 harvest). Recent research involving ABA and seed germination has shown mixed results, with Bruni (2001), finding there to be statistically significant biological effects, but Couchman (2001) not. Objective/Aim/Purpose The purpose of this study was to evaluate the effectiveness of homoeopathic dilutions of ABA, molybdenum and allopurinol (two substances which have an effect on ABA metabolism), especially those above the 10-23 level (Avogadro’s dilution limit), on germination, in light of recent findings. Abscisic acid, a plant hormone and molybdenum, a trace element, both play an essential role in inducing dormancy of the seed. Allopurinol, a therapeutic drug, has also been shown to affect ABA metabolism and therefore seed germination. The study used all three substances individually and in combination, in homoeopathic dilutions ranging from 4CH to 200CH potency. Methodology There were 7 treatments with 5 potencies per treatment (4CH, 9CH, 15CH, 30CH and 200CH). Each potency level for each treatment had a control, which meant there were 5 controls per treatment. The seeds (distally cut) were placed in 9cm Petri dishes (20 seeds in each), with 5 repetitions, 100 seeds per dilution level with one control of 20 seeds. There were thus 600 (120 x 5) seeds per treatment and 4200 seeds in total (600 x 7 treatments). Seeds were germinated in the dark at a constant temperature. Counts were done every 24 hours for 3 days and the data recorded. The criterion for germination was radical emergence. Results The data was analysed statistically using Univariate Analysis of Variance (STATISTICA version 6). The results showed statistically significant interaction between treatments and potencies and a One-Way Anova was then used to analyse each treatment to determine the effectiveness of each potency. Statistically significant differences were noted between potencies for each treatment. From the results it was clear that the most effective treatment for stimulating germination was the treatment utilizing homoeopathic dilutions of allopurinol. The most effective treatment for inhibiting germination was the treatment utilizing ABA in homoeopathic dilutions. The 30CH (10-60) showed a statistically significant effect on the stimulation of germination across almost all treatments, whereas the 15CH (10-30) showed a statistically significant effect in inhibiting germination in most treatments. Conclusion It is evident from the results of this study that all the treatments produced distinct biological effects, whether it be stimulating germination or inhibiting germination in homoeopathic dilution.

<p>Promoter detection, especially in prokaryotes, has always been an uphill task and may remain so, because of the many varieties of sigma factors employed by various organisms in transcription. The situation is made more complex by the fact, that any seemingly unimportant sequence segment may be turned into a promoter sequence by an activator or repressor (if the actual promoter sequence is made unavailable). Nevertheless, a computational approach to promoter detection has to be performed due to number of reasons. The obvious that comes to mind is the long and tedious process involved in elucidating promoters in the &lsquo / wet&rsquo / laboratories not to mention the financial aspect of such endeavors. Promoter detection/prediction of an organism with few characterized promoters (M.tuberculosis) as envisaged at the beginning of this work was never going to be easy. Even for the few known Mycobacterial promoters, most of the respective sigma factors associated with their transcription were not known. If the information (promoter-sigma) were available, the research would have been focused on categorizing the promoters according to sigma factors and training the methods on the respective categories. That is assuming that, there would be enough training data for the respective categories. Most promoter detection/prediction studies have been carried out on E.coli because of the availability of a number of experimentally characterized promoters (+- 310). Even then, no researcher to date has extended the research to the entire E.coli genome.</p>

Many aspects of biology depend on the ability of DNA-binding proteins to locate specific binding sites within the genome. This search process is required at the beginning of all site-specific protein-DNA interactions, and has the potential to act as the first stage of biological regulation. Given the difficulty of pinpointing a small region of DNA, within even simple genomes, it is expected that proteins are adapted to use specialized mechanisms, collectively referred to as facilitated diffusion [Berg et al., 1981], to effectively reduce the dimensionality of their searches, and rapidly find their targets. Here, we use a combination of nanofabricated microfluidic devices and single-molecule microscopy to determine whether facilitated diffusion contributes to all DNA target searches. We investigate promoter binding by E. coli RNA polymerase, foreign DNA recognition by CRISPR-Cas complexes, and Rad51’s homology search during recombination. In each example, we observe that the target searches proceed without extensive use of facilitated diffusion; rather, consideration of these non-facilitated target searches reveals an alternative search strategy. We show that instead of reducing the dimensionality of their searches, these proteins, reduce search complexity by minimizing unproductive interactions with DNA, thereby increase the probability of locating a specific DNA target.

Thesis (Ph. D.)--Ohio State University, 2003. / Title from first page of PDF file. Document formatted into pages; contains xii, 152 p. : ill., (some col.). Includes abstract and vita. Advisor: Anthony Young, Molecular, Cellular, and Developmental Biology Program. Includes bibliographical references (p. 137-150).

Triticale (x Triticosecale Whitm.) is a cereal with favorable agronomic traits for a Canadian bioproduction platform crop. Appropriate tissue sampling times were determined and gene expression profiles were evaluated in five triticale seed tissues and eleven vegetative tissues using the Affymetrix Wheat GeneChip®. Genes that were expressed, not expressed, tissue-specific, tissue-enriched and developmentally regulated were identified. The percentage of probe sets on the wheat GeneChip with gene ontology annotations was improved from less than 3% to over 76% using homologous sequence identification and annotation transfer. This information was used to determine functions and processes over-represented within the identified gene lists and provide biological meaning to the results. Expression of candidate genes was further evaluated using qRT-PCR, RNA in situ hybridization and promoter characterization. This study has provided a comprehensive triticale gene expression atlas; knowledge regarding triticale development, gene function, expression and regulation; and tools enabling further triticale research and development. / xxiii, 425 leaves : col. ill. ; 29 cm

<p>Promoter detection, especially in prokaryotes, has always been an uphill task and may remain so, because of the many varieties of sigma factors employed by various organisms in transcription. The situation is made more complex by the fact, that any seemingly unimportant sequence segment may be turned into a promoter sequence by an activator or repressor (if the actual promoter sequence is made unavailable). Nevertheless, a computational approach to promoter detection has to be performed due to number of reasons. The obvious that comes to mind is the long and tedious process involved in elucidating promoters in the &lsquo / wet&rsquo / laboratories not to mention the financial aspect of such endeavors. Promoter detection/prediction of an organism with few characterized promoters (M.tuberculosis) as envisaged at the beginning of this work was never going to be easy. Even for the few known Mycobacterial promoters, most of the respective sigma factors associated with their transcription were not known. If the information (promoter-sigma) were available, the research would have been focused on categorizing the promoters according to sigma factors and training the methods on the respective categories. That is assuming that, there would be enough training data for the respective categories. Most promoter detection/prediction studies have been carried out on E.coli because of the availability of a number of experimentally characterized promoters (+- 310). Even then, no researcher to date has extended the research to the entire E.coli genome.</p>

Cell culture systems have provided many insights into eukaryotic gene expression and other biochemical mechanisms. Since the cell represents the smallest living unit of any organism it provides a desirable in vitro system, allowing biochemical studies without the complex physiology of an entire animal. However, processes involving intracellular mechanisms, such as development, aging or carcinogenis, eventually require the analysis of the intact organism. Transgenic animals are a very promising tool to approach questions of this magnitude. Fish in general and the zebrafish (Brachydanio rerio) in particular are an excellent model system for transgenic research, mainly due to their extramaternal fertilization and development and their short generation cycle throughout the year. The recent derivation of zebrafish cell lines has opened up possibilities for in vitro analysis of this popular model species, and expression of heterologous genes under the influence of promoter and other regulatory nucleic acid aequences. In contrast to mammalian expression systems, little nucleic acid sequences controlling gene expression in fish are known. Therefore we examined mammalian expression systems in fish cells in order to determine their efficiency quantitatively. Emphasis was given to zebrafish cultures with the goal of eventually injecting in vitro manipulated embryo cells into host embryos and thereby creating transgenic chimera. / Graduation date: 1993

Bibliography: leaves 167-176. / xvi, 176 leaves : ill. ; 30 cm. / Title page, contents and abstract only. The complete thesis in print form is available from the University Library. / The aim of this thesis is to characterise a bacterial expression system for recombinant production of proteins with relevance to industry. Recombinant protein expression under control of stationary-phase inducible promoters is characterised, and the expression levels are quantitatively compared with those under control of IPTG-inducible promoters. A number of bacterial expression systems are constructed using promoter::cat transcriptional fusions. / Thesis (Ph.D.)--University of Adelaide, Depts. of Microbiology and Immunology and Chemical Engineering, 1997

[Formulae and special characters can only be approximated. Please see the pdf version of this abstract for an accurate reproduction.] Tumor necrosis factor (TNF) is a pleiotropic cytokine that mediates a long list of immunological and pathophysiological processes. TNF is produced by a wide variety of cells including immune and non-immune cells, however in most cell types TNF is not expressed prior to stimulation. The function of TNF is mediated via its trimeric domain by binding to TNF receptors that are found on most types of cells, especially of the haematopoietic systems, hence transpiring its effects on a wide variety of cells and organ systems. The cytotoxic (apoptosis) and pro-inflammatory (differentiation, proliferation and activation) functions of TNF are protective but can also result in pathological or deleterious consequences. A biallelic G to A transition polymorphism in the promoter region of TNF at nucleotide position 308 from the transcription start site is suggested to be involved in differential transcriptional regulation of TNF expression. The high TNF producing 308A allele is associated with susceptibility to or worse outcome of many infectious diseases in addition to autoimmune and other pathophysiological conditions. A previous study in our laboratory observed a selective affinity towards the polymorphic 308A allele by an EMSA protein(s) complex, named E. Several other protein complexes were found along with complex E and one of them was identified as Sp1. The identification of complex E was unsuccessful but it was hypothesized to play a major role as transcriptional activator in 308A allele individuals hence transpiring its effect in various pathophysiological states. In this study, the EMSA complexes observed in the TNF promoter region between nucleotides 322 to 283, encompassing the 308 polymorphism, is characterized. EMSA using mutated oligonucleotides mapped the binding sites of complexes B, C, D and E. TRANSFAC database search in addition to previous work revealed the identity of complex C as Sp1 but the rest of complexes remained unknown. Moreover, in contrast to our previous study, the protein(s) in the complex E was found to preferentially bind 308G nucleotide hence posing as a transcriptional repressor, resulting in decreased production state of TNF in 308G allele individuals than 308A allele individuals. In order to characterize putative transcription factors binding to the promoter region, first the biochemical characteristics such as the effects of temperature, salts and cations on DNA binding ability of EMSA complexes were studied. EMSA complexes B, C, DI and E required cations, probably Zn+2, to bind DNA. By optimizing a technique that couples EMSA with SDS-PAGE, the molecular weight of C, DI and E was determined. A novel technique that couples EMSA with IEF determined the pI of complexes B, C, D, DI and E. Although a commonly used technique of identifying unknown DNA-binding protein of interest, Yeast One-Hybrid assay, did not identify complex E, the novel identification method involving chromatography, two-dimensional electrophoresis, EMSA, mass spectrometry and database interrogation successfully identified TNF EMSA complex E as transcription factor Ying Yang 1 (YY1). Supershift EMSA confirmed complex E as YY1. In addition, the supershift assay showed presence of Sp1 and Sp3 in complex C. Similarly, complex DI is identified as Sp3. The novel method in identifying DNA-binding proteins is particularly useful as this technique allows identification of protein seen in EMSA without the need of extensive identification process. YY1 binds to a 6 base pair sequence, 5? TTGAGG 3?, from nt 295 to 290 of TNF promoter. The loss of affinity in 308A allele is caused by transition of underlined G nucleotide to A. The determined and described molecular weight of YY1 in literature is 60 kDa while the theoretical weight is 45 kDa. Both the determined and theoretical pI of YY1 is 5.8. YY1 is a multifunctional transcription factor implicated in both positive and negative regulation of gene expression as well as in initiation of transcription. It is ubiquitously expressed in growing, differentiated, and growth-arrested cells. Although future experiment is yet to establish in vivo presence of YY1 in TNF promoter, our study so far provides convincing evidence that the putative transcription factor that has selective affinity towards 308G allele is indeed YY1.

Thesis (Ph.D.)--University of Western Australia, 2004.