La recombinaison homologue est un mécanisme de réparation de l’ADN extrêmement contrôlé et particulièrement chez les eucaryotes supérieurs. Dans les cellules méiotiques de ces derniers, où les cassures doubles brin de l’ADN sont programmées, les voies de crossing-over de la recombinaison homologue, qui génèrent de nouvelles combinaisons de gènes, sont restreintes. Dans les cellules somatiques, la recombinaison illégitime, qui assure majoritairement la réparation des cassures double brin de l’ADN, limite l’intégration ciblée du transgène par recombinaison homologue. Les entreprises de biotechnologie convoitent de maitriser la recombinaison homologue afin de contrôler d’une part le brassage génomique qui a lieu pendant la méiose, et d’autre part l’intégration du transgène dans le génome. Cette étude a porté sur le développement d’outils afin d’atteindre ces deux objectifs. Afin d’augmenter le brassage du génome, ayant lieu pendant la méiose, une version du promoteur OsDmc1b, active dans les cellules méiotiques, a été caractérisée chez le maïs. Des plantes sur-exprimant le gène ZmSpo11.1, sous contrôle de ce promoteur, ont ainsi été développées afin d’obtenir des lignées potentiellement hyper-recombinantes. Si la surexpression de ZmSpo11.1 permet effectivement d’augmenter le taux de crossing-over, il pourra être utilisé par les sélectionneurs afin d’accélérer l’introgression d’allèles d’intérêt dans des variétés élites. Concernant la mise en place d’une technique de ciblage de gène, deux stratégies, reposant sur l’utilisation de la méganucléase I-SceI, ont été testées. La démarche a nécessité trois éléments : un locus cible contenant le site de coupure I-SceI, une matrice de réparation et la séquence codant I-SceI (ou I-SceI::GR). La première stratégie, consistant à retransformer les lignées présentant le locus cible avec la matrice de réparation et I-SceI, ne semble pas exploitable car aucun évènement de ciblage de gène n’a été mis en évidence. La seconde stratégie, reposant sur l’assemblage des trois éléments par croisement, est beaucoup plus prometteuse. Malgré la faible activité d’I-SceI::GR, des évènements de recombinaison homologue ont été observés dans les tissus foliaires de certaines plantes. Du cal embryogène, développé à partir de ces dernières, a permis de régénérer des plantes présentant des évènements de ciblage de gène. Ces travaux ouvrent de nouvelles perspectives dans l’élaboration contrôlée d’OGM. / Homologous recombination is a DNA repair mechanism highly regulated in higher eukaryotes. In their meiotic cells, where DNA double-stranded breaks are programmed, the crossing-over pathway of homologous recombination, which generates new gene combinations, is limited in activity and genomic distribution. In somatic cells, illegitimate recombination, which mainly ensures DNA double-strand repair, limits the targeted integration of transgenes by homologous recombination. Biotechnology companies aim to master homologous recombination to control on the one hand the genomic mixing that occurs during meiosis, and on other hand, the integration of transgenes into the genome. This study focuses on the development of tools to achieve these two objectives.To increase genome mixing occurring during meiosis, a version of the OsDmc1b promoter active in maize meiotic cells was isolated. Then, plants over-expressing the ZmSpo11.1 gene under control of this promoter have been developed to obtain potentially hyper-recombinant lines. If ZmSPO11.1 overexpression increases the crossing over rate, it can be used by breeders to accelerate the introgression of alleles of interest into elite varieties. For the establishment of a gene targeting technique, two strategies based on the use of the I-SceI meganuclease were tested. These approaches involved the use of three elements which are: a target locus containing the cleavage site of I-SceI, a repair template and the sequence encoding I-SceI (or ISceI::GR). The first strategy, consisting of the retransformation of target locus lines with the repair template and I-SceI, does not seem workable because no gene targeting events were isolated. The second strategy, based on the assembly of the three components by crossing, is more promising. Despite the low activity of I-SceI::GR, homologous recombination events were observed in leaf tissues of certain plants. Embryogenic callus, developed from these plants, permitted the regeneration of plants with gene targeting events. This work opens new perspectives in the development of controlled GMO production.

The period of ice break-off in spring is a key event for many biogeochemical processes in lakes globallly. The biogeochemical processes occurring at ice break-off have the potential of influencing characteristics of lakes throughout spring and summer, including algal blooms and greenhouse gas emission. This makes it important to study lakes in the period of ice break-off. At ice break-off, soil bacteria from the catchment area usually enter the lake via spring floods and mix with the bacteria already occurring in the lake water. In this study, the effects of mixing soil- and lake microbial communities during ice break-off-like conditions were tested by performing an experiment under controlled conditions in the laboratory. In the experiment, light, microbial community composition and concentration of soil-derived organic matter were manipulated to simulate different conditions associated with ice break-off. The variables investigated were bacterial activity and functionality, measured as cell abundance and enzymatic activity, as well as primary production and concentration of dissolved organic matter. The results showed that a mix of soil and lake microbial communities had enzymatic activity patterns resembling lake communities, and then shifted to being more similar to soil communities. The experiment also showed that degradation of measured dissolved organic matter was not linked to biotic processes, and that the observed decrease was most likely due to photo degradation. Finally, the experiment showed that primary production, here measured as chlorophyll a, was only stimulated by the mixed community with light and added soil dissolved organic matter. The results found in this study are important as they show that microbial communities do alter their function and enzymatic activity based on composition. Furthermore, the result that primary production was only seen in the presence of light, soilderived organic matter and a mixed community of lake and soil bacteria may be seen as an indication that primary producers in lake ecosystems to some extent depend on the inflow of terrestrial microbes and organic matter. It also possible that the coalescence of microbial communities enables the communities to perform tasks they were unable to prior to coalescence (i.e. perform tasks that allows primary production to take place). These results give the basis for further, more detailed studies.

Orientador: Cláudia Valéria Seullner Brandão / Resumo: O objetivo deste estudo foi comparar e avaliar a aplicação de célula tronco mesenquimal de tecido adiposo (CTM-TA) e plasma rico em plaquetas aquecido alogênico (PRP-AA) perilacrimal em cães com ceratoconjuntivite seca (CCS), bem como, se há diferença na resposta segundo o grau de gravidade da afecção. Foram analisados 20 cães com produção lacrimal <15mm/min e distribuídos aleatoriamente em dois grupos (n=10). O grupo 1 tratado com CTM-TA (5x106 e 3x106 células) (GCTM-TA) e com grupo 2 PRP-AA (0,7mL e 0,3mL) (GPRP-AA), ambos injetados perilacrimal nas glândulas lacrimal principal e da terceira pálpebra, respectivamente. Todos os olhos foram avaliados em quatro momentos (M0 M15, M30, M60 dias). As variáveis avaliadas foram: osmolaridade da lágrima; teste lacrimal de Schirmer (TLS); sensibilidade corneal; tempo de ruptura do filme lacrimal (TRFL); pressão intraocular; espessura corneal; e biopsia da conjuntiva bulbar, além das variáveis clínico-oftalmológicas. Houve melhora nos dois grupos a partir de M15 (p<0,05) na qualidade do filme lacrimal, avaliada por meio da osmolaridade e TRFL, sem diferença entre os grupos. A produção de lágrima aumentou, entretanto notou-se diferença significativa nos animais com CCS discreta no GPRP-AA a partir do M30, e nos cães com CCS grave no GCTM-TA a partir do M30 e no GPRP-AA a partir do M60. Verificou-se melhora dos sinais clínicos da inflamação em ambos os grupos. Aplicação única perilacrimal de CTM-TA e PRP aquecido alogênicos normaliza a ... (Resumo completo, clicar acesso eletrônico abaixo) / Abstract: The aim of the study was to compare and evaluate the application of allogenic mesenchymal stem cell adipose tissue derived (MSCs-AD) and inativate platelet-rich plasma allogenic (PRP-IA), perilacrimal in dogs with keratoconjunctivitis sicca (KCS), and if there is difference in the response according to the degree of severity of the condition. Twenty dogs with lacrimal production <15mm/min randomly assigned to two groups were used. Group treated with MSCs-AD (5x106 and 3x106) (GMSCs-AD) and with PRP-IA (0.7ml and 0.3ml) (GPRP-IA), both intralacrimal in the main lacrimal glands and third eyelid, respectively. The eyes were evaluated in four moments after application (M0 M15, M30, M60). The variables evaluated were: tear osmolarity e; Schirmer's tear test (STT); sensitivity corneal; tear film break time (TBUT); intraocular pressure; corneal thickness; and biopsy of the bulbar conjunctiva, in addition to clinical-ophthalmologic variables. There was improvement in two groups at M15 (p <0.05) on tear film quality, assessed by osmolarity and TBUT, with there was no difference between groups. STT increased, however, statistical difference was only observed in animals with mild KCS in the GPRP-IA from the M30 and in the intense KCS at M30 in the G MSCs-AD and at M60 in the GPRP-IA. There was an improvement in the clinical signs of inflammation in both groups. The unique Intralacrimal application of MSCs-AD and PRP-IA normalizes tear film quality and improves clinical signs of inflamma... (Complete abstract click electronic access below) / Doutor

In order to keep shared access computers secure and stable system administrators resort to locking down the computing environment in order to prevent intentional and unintentional damage by users. Skilled attackers are often able to break out of locked down computing environments and intentionally misuse shared access computers. This misuse has resulted in cases of mass identity theft and fraud, some of which have had an estimated cost ranging in millions. In order to determine if it is possible to break out of locked down computing environments an assessment method is required. Although a number of vulnerability assessment techniques exist, none of the existing techniques are sufficient for assessing locked down shared access computers. This is due to the existing techniques focusing on traditional, application specific, software vulnerabilities. Break out path vulnerabilities (which are exploited by attackers in order to break out of locked down environments) differ substantially from traditional vulnerabilities, and as a consequence are not easily discovered using existing techniques. Ontologies can be thought of as a modelling technique that can be used to capture expert knowledge about a domain of interest. The method for discovering break out paths in locked down computers can be considered expert knowledge in the domain of shared access computer security. This research proposes an ontology based assessment process for discovering break out path vulnerabilities in locked down shared access computers. The proposed approach is called the ontological lockdown assessment process. The ontological lockdown assessment process is implemented against a real world system and successfully identifies numerous break out path vulnerabilities.

PhD / SYNOPSIS OF THIS THESIS The aim of this thesis is to more fully understand and explain the binding mechanism of organic molecules to the Au(111) surface and to explore the conduction of such molecules. It consists of five discreet chapters connected to each other by the central theme of “The Single Molecule Device: Conductance and Binding”. There is a deliberate concentration on azine linkers, in particular those with a 1,10-phenanthroline-type bidentate configuration at each end. This linker unit is called a “molecular alligator clip” and is investigated as an alternative to the thiol linker unit more commonly used. Chapter 1 places the work in the broad context of Molecular Electronics and establishes the need for this research. In Chapter 2 the multiple break-junction technique (using a Scanning Tunnelling Microscope or similar device) was used to investigate the conductance of various molecules with azine linkers. A major finding of those experiments is that solvent interactions are a key factor in the conductance signal of particular molecules. Some solvents interfere with the molecule’s interaction with and attachment to the gold electrodes. One indicator of the degree of this interference is the extent of the enhancement or otherwise of the gold quantized conduction peak at 1.0 G0. Below 1.0 G0 a broad range for which the molecule enhances conduction indicates that solvent interactions contribute to a variety of structures which could bridge the electrodes, each with their own specific conductance value. The use of histograms with a Log10 scale for conductance proved useful for observing broad range features. vi Another factor which affects the conductance signal is the geometric alignment of the molecule (or the molecule-solvent structure) to the gold electrode, and the molecular alignment is explored in Chapters 3 for 1,10-phenanthroline (PHEN) and Chapter 4 for thiols. In Chapter 3 STM images, electrochemistry, and Density Functional Theory (DFT) are used to determine 1,10-phenanthroline (PHEN) structures on the Au(111) surface. It is established that PHEN binds in two modes, a physisorbed state and a chemisorbed state. The chemisorbed state is more stable and involves the extraction of gold from the bulk to form adatom-PHEN entities which are highly mobile on the gold surface. Surface pitting is viewed as evidential of the formation of the adatom-molecule entities. DFT calculations in this chapter were performed by Ante Bilic and Jeffery Reimers. The conclusions to Chapter 3 implicate the adatom as a binding mode of thiols to gold and this is explored in Chapter 4 by a timely review of nascent research in the field. The adatom motif is identified as the major binding structure for thiol terminated molecules to gold, using the explanation of surface pitting in Chapter 3 as major evidence and substantiated by emergent literature, both experimental and theoretical. Furthermore, the effect of this binding mode on conductance is explored and structures relevant to the break-junction experiment of Chapter 2 are identified and their conductance values compared. Finally, as a result of researching extensive reports of molecular conductance values, and having attempted the same, a simple method for predicting the conductance of single molecules is presented based upon the tunneling conductance formula.

<p>Sammanfattning</p><p>I en pappersrulle är de yttre lagren utsatta för dragspänning. Om det finns variationer i rullens diameter i form av valkar finns det risk för att papperet förlängs irreversibelt i dessa positioner och orsaka slappa stråk. Den deformation som uppstår vid långvarig belastning av papper kallas krypning.</p><p>Uppgiften i detta arbete var, att på lab., simulera valkar genom att utsätta pappersremsor för konstant dragspänning av olika storlekar. En av frågeställningarna var om egenskapsförändringar kan påvisas när papper utsätts för konstant inspänningskraft under en längre tid. De parametrar som undersökts var töjning, kryphastighet och relaxation, även mätningar på papperets mekaniska egenskaper dragstyrka, brottöjning och dragstyvhet har utförts i detta arbete.</p><p>De papperskvalitéer som undersöktes var 71g/m2 MG-papper, 80g/m2 optisk liner och 135g/m2 liner. Inspänningskraften som papperet utsattes för motsvarade 10 %, 30 % och 50 % av dragstyrkan hos varje kvalité. Provremsor belastades under ett, fyra och sju dygn, för att sedan relaxeras.</p><p>Resultaten visar att dragstyrkan och dragstyvheten inte påverkas av att papperet varit utsatt för en inspänningskraft. Däremot blev brottöjningen mindre ju större den irreversibla förlängningen hos papperet var. Samtliga papperskvalitéer relaxerar tillbaka till ursprungslängd efter fyra dygn när belastningen var 10 %. Om belastningen däremot var 30 % och 50 % uppstod en irreversibel förlängning redan efter ett dygn. Vid belastning under sju dygn uppstod permanent förlängning hos samtliga kvalitéer och belastningar utom MG-papper 10 % som relaxerade till ursprungslängd. MG-papper är därmed den papperskvalité som står emot krypning bäst och är på så sätt minst känslig för valkar i pappersrullen.</p><p>En högre belastning ger en högre kryphastighet och större irreversibel förlängning. För att undvika uppkomsten av slappa stråk bör därför banspänningen som används vid upprullningen av papperet inte vara större än 10 % av dragstyrkan hos papperskvalitén.</p><p>Hur länge papperet är belastat påverkar också den slutliga irreversibla förlängningen. Det innebär att lagring ökar risken för att slappa stråk kommer att uppstå i efterbearbetningen.</p> / <p>Abstract</p><p>In a paper roll the outer layers are under tensile stress. If there are variations in the roll diameter there is a risk that the thicker parts of the roll will make the paper elongate more in these regions, because they will be under a higher tensile stress. This may cause baggy webs. The deformation that occurs during a long time under tensile stress is called creep.</p><p>Experiments have been performed with strips of paper that have been exposed to a constant load and constant climate (50 % RH, 23°C). The properties that have been examined are creep strain, creep rate and relaxation after creep. Studies of tensile strength, tensile stiffness and strain at break have also been made.</p><p>The paper qualities that have been examined were 71g/m2 MG-paper, 80g/m2 liner and 135g/m2 liner. The paper qualities in this study are exposed to loads that correspond to 10 %, 30 % and 50 % of their tensile strength. The paper strips were under load during 24h, four and seven days.</p><p>The results in these studies show that tensile strength and tensile stiffness remains the same after the paper has been exposed to a constant load. The strain at break, on the other hand, decreases with increasing irreversible elongation of the paper strips. All paper qualities recover to the initial length after four days under a load of 10 % of the tensile strength. If the load corresponds to 30 % or 50 % of the tensile strength a permanent elongation arose as early as after 24h. After seven days, only the MG-paper recovered to the initial length if the load corresponded to 10 % of the tensile strength. This means that MG-paper is more resistant to creep then the liner qualities. Consequently, MG-paper is the paper quality that is least sensitive to ridges.</p><p>A higher load gives a higher creep rate and larger irreversible elongation of the paper. To avoid baggy webs to arise, the load should not exceed 10 % of the paper’s tensile strength when winding the paper roll. The time under which the paper is under load also influence the final irreversible elongation. Therefore, to avoid baggy webs the paper roll shouldn’t be stored for too long.</p>

Thesis (M.Ing. (Development and Management))--North-West University, Potchefstroom Campus, 2007.

Thesis (M.Eng. (Development and Management Engineering)--North-West University, Potchefstroom Campus, 2008.

A two-dimensional numerical model (RiverFLO-2D) has been enhanced to simulate flooding of urban areas by developing an innovative wet and dry surface algorithm, accounting for variable rainfall, and recoding the model computer program for parallel computing. The model formulation is based on the shallow water equations solved with an explicit time-stepping element-by-element finite element method. The dry-wet surface algorithm is based on a local approximation of the continuity and momentum equations for elements that are completely dry. This algorithm achieves global volume conservation in the finite element, even for flows over complex topographic surfaces. A new module was implemented to account for variable rainfall in space and time using NEXRAD precipitation estimates. The resulting computer code was parallelized using OpenMP Application Program Interface, which allows the model to run up to 5 times faster on multiple core computers. The model was verified with analytical solutions and validated with laboratory and field data. Model application to the Malpasset dam break and Sumacarcel flooding event show that the model accurately predicts flood wave travel times and water depths for these numerically demanding real cases. To illustrate the predictive capability of the enhanced model, an application was made of the city of Sweetwater flooding in Miami-Dade County, FL caused by the Hurricane Irene. The simulation starts with dry bed and rainfall is provided by NEXRAD estimates. Integrating NEXRAD rainfall estimates, developing a novel dry-wet area algorithm and parallelizing RiverFLO-2D code, this dissertation presents a proof of concept to accurately and efficiently predict floods in urban areas, identifying future improvements along this line of research.

Investigations of DNA damage response (DDR) mechanisms in normal tissues have implications for both cancer prevention and treatments. The accumulating knowledge about protein function and molecular markers makes it possible to directly trace and interpret cellular DDR in a tissue context. Using immunohistochemical techniques and digital image analysis, we have examined several principal DDR events in epidermis from patients undergoing fractionated radiotherapy. Acquiring biopsies from different regions of the skin provides the possibility to determine in vivo dose response at clinically relevant dose levels throughout the treatment. A crucial event in cellular DDR is the repair of DNA double strand breaks (DSBs). These serious lesions can be directly visualised in cells by detecting foci forming markers such as γH2AX and 53BP1. Our results reveal that DSB-signalling foci can be detected and quantified in paraffin-embedded tissues. More importantly, epidermal DSB foci dose response reveals hypersensitivity, detected as elevated foci levels per dose unit, for doses below ~0.3Gy. The low-dose hypersensitive dose response is observed throughout the treatment course and also in between fractions: at 30 minutes, 3 hours and 24 hours following delivered fractions. The dose response at 24 hours further reveals that foci levels do not return to background levels between fractions. Furthermore, a low-dose hypersensitive dose response is also observed for these persistent foci. Investigations of end points further downstream in the DDR pathways confirmed that the low-dose hypersensitivity was preserved for: the checkpoint regulating p21 kinase inhibitor; mitosis suppression; apoptosis induction and basal keratinocyte reduction. Our results reveal preserved low-dose hypersensitivity both early and late in the DDR pathways. A possible link between the dose-response relationships is therefore suggested. The preserved low-dose hypersensitivity is a cause for re-evaluation of the risks associated with low-dose exposure and has implications for cancer treatments, diagnostics and radiation protection.