Global warming has been a topic of discussion since the discovery that man-made greenhouse gas emissions is having an affect on the planet almost 50 years ago. Grocery products stand for roughly one third of all EUs carbon emissions as a result of its highvolume production. Carbon labeling is a tool in which retailers and manufacturers can communicate the amount has caused throughout its whole life cycle or that the they are working towards lowering their GHG emission throughout their organization. However, previous research indicates that carbon labels has not yet had its breakthrough moment yet due to the CO2e data is too complex for the consumers to interpret. This led to our research question “What is the preferred content, design and layout of a carbon label among Swedish consumers? “The purpose of this study is to get a deeper understanding into the preferences of the Swedish consumers and what kind of attributes they are looking for to make a carbon label understandable. We found that the Swedish consumer prefers a more complex design than previous studies have suggested based on researches made from other countries. In order to do our explanatory research, we measured these variables with a quantitative survey and made statistical calculations such as mean values and correlation analysis to see if our hypotheses were supported.  The analysis shows that the Swedish consumers prefer all the following attributes that is being presented in an order of priority: The label should be colour coded, made by a well-known organization, presented in terms of scale and have the CO2e data presented on the label.

Global threats to treatment of bacterial infections due to antibiotic resistance (AR) have been on the rise in recent years. Current diagnostic tests identify bacteria by using blood culture, which takes more than 24 hours. This study focuses on the fluorescent labeling of DNA derived from bacterial AR genes (KPC & VIM) and other model DNAs using oligreen dye (OG) and molecular beacons (MB). A NanoDrop 3300 fluorospectrometer was used to take fluorescence measurements. Linear dynamic range and labeling efficiency were dependent on the following optimized conditions: dilution factor of OG (200 fold), buffer (20 mM Tris HCl, pH 8), and heat treatment of 95 °C for 15 min.Fluorescence analysis of a target DNA with a designed MB showed signal-to-background of 10 with our buffer only and 20 with our buffer and 25% ethanol. I also demonstrated a simple microfluidic device capable of detecting AR genes using model DNAs, magnetic beads, and designed MBs for assays of µ50 L volume. This study provides a first step towards detecting MB-DNA complexes by a simple, low cost, and fast non-amplified method, which may be used to detect AR genes in clinical samples in the future.

L'étude de la dynamique des protéines est essentielle pour comprendre les processus biologiques. Notre laboratoire a développé une nouvelle classe de protéines fluorescentes semi-synthétiques, appelée Fluorescence-Activating and absorption-Shifting Tag (FAST). Cette thèse de doctorat présente le développement de nouveaux systèmes FAST avec diverses propriétés pour l'imagerie multiplexée. Nous avons développé une série de fluorogènes permettant de modifier la couleur de FAST de vert-jaune à orange et rouge. Au delà de l’application de l’imagerie multi-couleurs, ces fluorogènes permettant un échange dynamique des couleurs grâce à la liaison réversible de FAST, ouvrant de nouvelles perspectives pour le développement de méthodes d’imagerie sélective reposant sur la dynamique de systèmes réactifs. Pour étendre davantage les propriétés spectrales de FAST vers le rouge lointain, nous avons développé une nouvelle série de fluorogènes rouges, pour lesquels nous avons sélectionné par une stratégie d'évolution dirigée basée sur le yeast display et la cytométrie en flux de nouveaux tags protéiques capables d’interagir avec ces fluorogènes et d’activer leur fluorescence. Nous avons enfin développé de nouveaux fluorogènes capables de former des complexes fluorescents avec FAST, mais incapables de traverser la membrane plasmique, ce qui permet de détecter sélectivement les protéines membranaires. / Studying protein activities could help us to understand the complex mechanisms controlling cells and organisms. Our laboratory recently developed Fluorescence-Activating and absorption-Shifting Tag (FAST), a small fluorogen-based reporter enabling to fluorescently label fusion proteins in living cells. My PhD thesis presents the developments of new FAST systems with various properties for multiplexed imaging. We report a collection of fluorogens enabling to tune the fluorescence color of FAST from green-yellow to orange and red. Beyond allowing multicolor imaging of FAST-tagged proteins in live cells, these fluorogens enable dynamic color switching because of FAST’s reversible labeling, opening great prospects for the design of selective imaging methods relying on dynamic systems. In order to further expand the spectral properties of FAST to red, we also designed and developed a library of red fluorogenic dyes, for which we engineered specific protein binders by applying a directed evolution strategy based on the yeast display technology and high-throughput fluorescence activating cell sorting (FACS). We finally developed novel fluorogens able to form fluorescent complexes with FAST, but incapable of crossing the plasma membrane, which makes it possible to selectively detect FAST-tagged cell-surface proteins.

For k∈{Z}+ and G a simple connected graph, a k-radio labeling f:VG→Z+ of G requires all pairs of distinct vertices u and v to satisfy |f(u)-f(v)|≥ k+1-d(u,v). When k=1, this requirement gives rise to the familiar labeling known as vertex coloring for which each vertex of a graph is labeled so that adjacent vertices have different "colors". We consider k-radio labelings of G when k=diam(G). In this setting, no two vertices can have the same label, so graphs that have radio labelings of consecutive integers are one extreme on the spectrum of possibilities; graphs that can be labeled with such a labeling are called radio graceful. In this thesis, we give four main results on the existence of radio graceful graphs, which focus on Hamming graphs (Cartesian products of complete graphs) and a generalization of the Petersen graph. In particular, we prove the existence of radio graceful graphs of arbitrary diameter, a result previously unknown. Two of these main results show that, under certain conditions, the tth Cartesian power Gt of a radio graceful graph G is also radio graceful. We will also speak to occasions when Gt is not radio graceful despite G being so, as well as some partial results about necessary and sufficient conditions for a graph G so that Gt is radio graceful.

Ces travaux de thèse, dans le domaine de l'adéquation algorithme architecture pour la vision par ordinateur, ont pour cadre l'étiquetage en composantes connexes (ECC) dans le contexte parallèle des architectures hautes performances. Alors que les architectures généralistes modernes sont multi-coeur, les algorithmes d'ECC sont majoritairement séquentiels, irréguliers et utilisent une structure de graphe pour représenter les relations d'équivalences entre étiquettes ce qui rend complexe leur parallélisation. L'ECC permet à partir d'une image binaire, de regrouper sous une même étiquette tous les pixels connexes, il fait ainsi le pont entre les traitements bas niveaux tels que le filtrage et ceux de haut niveau tels que la reconnaissance de forme ou la prise de décision. Il est donc impliqué dans un grand nombre de chaînes de traitements qui nécessitent l'analyse d'image segmentées. L'accélération de cette étape représente donc un enjeu pour tout un ensemble d'algorithmes.Les travaux de thèse se sont tout d'abord concentrés sur les performances comparées des algorithmes de l'état de l'art tant pour l'ECC que pour l'analyse des caractéristiques des composantes connexes (ACC) afin d'en dégager une hiérarchie et d’identifier les composantes déterminantes des algorithmes. Pour cela, une méthode d'évaluation des performances, reproductible et indépendante du domaine applicatif, a été proposée et appliquée à un ensemble représentatif des algorithmes de l'état de l'art. Les résultats montrent que l'algorithme séquentiel le plus rapide est l'algorithme LSL qui manipule des segments contrairement aux autres algorithmes qui manipulent des pixels.Dans un deuxième temps, une méthode de parallélisation des algorithmes directs utilisant OpenMP a été proposé avec pour objectif principal de réaliser l’ACC à la volée et de diminuer le coût de la communication entre les threads. Pour cela, l'image binaire est découpée en bandes traitées en parallèle sur chaque coeur du l'architecture, puis une étape de fusion pyramidale d'ensembles deux à deux disjoint d'étiquettes permet d'obtenir l'image complètement étiquetée sans avoir de concurrence d'accès aux données entre les différents threads. La procédure d'évaluation des performances appliquée a des machines de degré de parallélisme variés, a démontré que la méthode de parallélisation proposée était efficace et qu'elle s'appliquait à tous les algorithmes directs. L'algorithme LSL s'est encore avéré être le plus rapide et le seul adapté à l'augmentation du nombre de coeurs du fait de son approche «segments». Pour une architecture à 60 coeurs, l'algorithme LSL permet de traiter de 42,4 milliards de pixels par seconde pour des images de taille 8192x8192, tandis que le plus rapide des algorithmes pixels est limité par la bande passante et sature à 5,8 milliards de pixels par seconde.Après ces travaux, notre attention s'est portée sur les algorithmes d'ECC itératifs dans le but de développer des algorithmes pour les architectures manycore et GPU. Les algorithmes itératifs se basant sur un mécanisme de propagation des étiquettes de proche en proche, aucune autre structure que l'image n'est nécessaire ce qui permet d'en réaliser une implémentation massivement parallèle (MPAR). Ces travaux ont menés à la création de deux nouveaux algorithmes.- Une amélioration incrémentale de MPAR utilisant un ensemble de mécanismes tels qu'un balayage alternatif, l'utilisation d'instructions SIMD ainsi qu'un mécanisme de tuiles actives permettant de répartir la charge entre les différents coeurs tout en limitant le traitement des pixels aux zones actives de l'image et à leurs voisines.- Un algorithme mettant en œuvre la relation d’équivalence directement dans l’image pour réduire le nombre d'itérations nécessaires à l'étiquetage. Une implémentation pour GPU basée sur les instructions atomic avec un pré-étiquetage en mémoire locale a été réalisée et s'est révélée efficace dès les images de petite taille. / This PHD work take place in the field of algorithm-architecture matching for computer vision, specifically for the connected component labeling (CCL) for high performance parallel architectures.While modern architectures are overwhelmingly multi-core, CCL algorithms are mostly sequential, irregular and they use a graph structure to represent the equivalences between labels. This aspects make their parallelization challenging.CCL processes a binary image and gathers under the same label all the connected pixels, doing so CCL is a bridge between low level operations like filtering and high level ones like shape recognition and decision-making.It is involved in a large number of processing chains that require segmented image analysis. The acceleration of this step is therefore an issue for a variety of algorithms.At first, the PHD work focused on the comparative performance of the State-of-the-Art algorithms, as for CCL than for the features analysis of the connected components (CCA) in order to identify a hierarchy and the critical components of the algorithms. For this, a benchmarking method, reproducible and independent of the application domain was proposed and applied to a representative set of State-of-the-Art algorithms. The results show that the fastest sequential algorithm is the LSL algorithm which manipulates segments unlike other algorithms that manipulate pixels.Secondly, a parallelization framework of directs algorithms based on OpenMP was proposed with the main objective to compute the CCA on the fly and reduce the cost of communication between threads.For this, the binary image is divided into bands processed in parallel on each core of the architecture and a pyramidal fusion step that processes the generated disjoint sets of labels provides the fully labeled image without concurrent access to data between threads.The benchmarking procedure applied to several machines of various parallelism level, shows that the proposed parallelization framework applies to all the direct algorithms.The LSL algorithm is once again the fastest and the only one suitable when the number of cores increases due to its run-based conception. With an architecture of 60 cores, the LSL algorithm can process 42.4 billion pixels per second for images of 8192x8192 pixels, while the fastest pixel-based algorithm is limited by the bandwidth and saturates at 5.8 billion pixels per second.After these works, our attention focused on iterative CCL algorithms in order to develop new algorithms for many-core and GPU architectures. The Iterative algorithms are based on a local propagation mechanism without supplementary equivalence structure which allows to achieve a massively parallel implementation (MPAR). This work led to the creation of two new algorithms.- An incremental improvement of MPAR using a set of mechanisms such as an alternative scanning, the use of SIMD instructions and an active tile mechanism to distribute the load between the different cores while limiting the processing of the pixels to the active areas of the image and to their neighbors.- An algorithm that implements the equivalence relation directly into the image to reduce the number of iterations required for labeling. An implementation for GPU, based on atomic instructions with a pre-labeling in the local memory has been realized and it has proven effective from the small images.

The synthesis of proteins bearing unnatural amino acids has the potential to enhance and elucidate many processes in biochemistry and molecular biology. There are two primary methods for site specific unnatural amino acid incorporation, both of which use the cell’s native protein translating machinery: in vitro chemical acylation of suppressor tRNAs and the use of orthogonal amino acyl tRNA synthetases. Total chemical synthesis is theoretically possible, but current methods severely limit the maximum size of the product protein. In vivo orthogonal synthetase methods suffer from the high cost of the unnatural amino acid. In this thesis I sought to address this limitation by increasing cell density, first in shake flasks and then in a bioreactor in order to increase the yield of protein per amount of unnatural amino acid used. In a parallel project, I used the in vitro chemical acylation system to incorporate several unnatural amino acids, key among them the fluorophore BODIPYFL, with the aim of producing site specifically fluorescently labeled protein for single molecule FRET studies. I demonstrated successful incorporation of these amino acids into the trial protein GFP, although incorporation was not demonstrated in the final target, FEN1. This also served to confirm the effectiveness of a new procedure developed for chemical acylation.

京都大学 / 新制・課程博士 / 博士(理学) / 甲第23031号 / 理博第4708号 / 新制||理||1675(附属図書館) / 京都大学大学院理学研究科化学専攻 / (主査)教授 寺嶋 正秀, 教授 林 重彦, 教授 渡邊 一也 / 学位規則第4条第1項該当 / Doctor of Science / Kyoto University / DGAM

The aim of this study is to examine what people with a late (adult) ADHD diagnosis experience was contributing to the late diagnosis. The empirism of this study is mainly interviews with adults that have been diagnosed with ADHD. By applying a thematic analysisas a method, the empirics have been able to be categorized and analyzed. Based on the theoretical models of the labeling theory and locus of control, the result has been interpreted as meaning that the interviewees were labeled incorrectly before diagnosis took place and that a big development can be seen around the person's locus of control. The result shows that there are numerous factors which can be claimed to contribute to a late diagnosis. The majority of the respondents highlight the importance of the environment and mostly adults whether its school staff or members of the family. Factors such as cultural norms, prejudices and lack of knowledge are also crucial when it comes to when in life one gets his diagnosis. The study also examines how it affects people to recieve a late diagnosis, the result shows that an early diagnosis would have reduced the respondents sufferings and improve their life quality. / Studiens syfte är att undersöka vad personer med en sen (vuxen) ADHD-diagnos upplever bidrog till den sena diagnosen. Empirin av denna studie består främst av intervjupersoner med vuxna som har diagnostiserats med ADHD. Genom att tillämpa en tematisk innehållsanalys som metod har empirin kunnat kategoriseras och analyseras. Utifrån de teoretiska modellerna om stämplingsteorin och kontrollfokus har resultatet kunnat tolkas som att intervjupersonerna stämplats felaktigt innan diagnostisering skett samt att man kan se en rejäl utveckling kring personens locus of control. Resultatet visar att det finns flera faktorer som kan påstås bidra till en sen diagnos. Majoriteten av respondenterna belyser omgivnings betydelse och främst vuxna vare sig det är skolans personal eller familjemedlemmar. Faktorer såsom kulturella normer, fördomar och brist på kunskap är också avgörande för när individen får sin diagnos. Studien syftar också till att undersöka vilken effekt det har på respondenterna att få en sen diagnos, resultat visar att en tidig diagnos hade reducerat respondenternas lidande och förbättrat deras livskvalité.

<p>Plants naturally deposit a significant amount of carbon towards lignin, a polymer that imparts mechanical strength to cell walls but impedes our utilization of the polysaccharides in lignocellulosic biomass. Genetic engineering of lignin has demonstrated profound success in improving the processing of the biomass. Lignin is derived from the phenylpropanoid pathway, the architecture of which is well understood based upon the biochemical and genetic studies conducted to date. In contrast, we lack a systematic and quantitative view of the factors that determine carbon flux into and within this branched metabolic pathway in plants. To explore the control of carbon allocation for phenylalanine and lignin biosynthesis, we have developed a kinetic model of the pathway in Arabidopsis to test the regulatory role of several key enzymatic steps. We first established a ¹³C isotope feeding system for the measurement of flux using excised wild-type Arabidopsis stems. The excised stems continued to grow and lignify in our feeding system. When ring ¹³C₆-labeled phenylalanine ([¹³C₆]-Phe) was supplied to excised stems, isotope label was rapidly incorporated into soluble intermediates and lignin. Using this approach, we then analyzed metabolite pool sizes and isotope abundances of the pathway intermediates in a time course from stems fed with [¹³C₆]-Phe of different concentrations, and used these data to parameterize a kinetic model constructed with Michaelis-Menten kinetics. Our model of the general phenylpropanoid pathway captured the dynamic trends of metabolite pools <i>in vivo</i>and predicted the metabolic profiles of an independent feeding experiment. Based on the model simulation, we found that subcellular sequestration of pathway intermediates is necessary to maintain lignification homeostasis when metabolites are over-accumulated. Both the measurements and simulation suggested that theavailability of substrate Phe is one limiting factor for lignin flux in developing stems. This finding indicates new gene targets for lignin manipulation in plants. To extend our kinetic model to simulate flux distribution in response to genetic perturbations, we conducted an RNA-sequencing experiment in wild type and 13 plants with modified lignification, and integrated the transcriptional data with the metabolic profiles. We found that the biosynthesis of Phe and lignification are tightly coordinated at transcriptional level. The coregulation of the shikimate and phenylpropanoid pathways involves transcriptional and post-translational regulatory mechanisms to maintain pathway homeostasis. Our results also indicate that induction of Phe supply and enhancement of PAL activity are both effective strategies to increase carbon flux into the phenylpropanoid network.</p><p>In this interdisciplinary project, we have taken various system biology approaches to understand metabolic flux towards lignin, the second most abundant carbon sink in nature. We have combined isotope labeling aided flux measurements and mathematical simulation, and have integrated metabolome data with transcriptome profiles. The experiments and analysis have been conducted in both wild-type Arabidopsis and those with perturbed lignification. The novel work not only provides insight into our knowledge of phenylpropanoid metabolism, but also creates a framework to systematically assemble gene expression, enzyme activity, and metabolite accumulation to study metabolic fluxes, the ultimate functional phenotypes of biochemical networks.</p>

(IN ENGLISH) Structural mass spectrometry (MS) is an evolving field of structural biology introducing novel techniques for the characterization of biomolecules. Although MS-based techniques only can provide "low-resolution" information compared to standard high-resolution techniques representing by X-ray crystallography, cryo-electron microscopy or nuclear magnetic resonance, its uniqueness lies in the ability to easily obtain structural information about various biomolecules in their native or native-like environment. By employing various approaches, from protein covalent labelling through chemical cross- linking to ion mobility, structural MS provides insight into the structure and dynamics of proteins and their complexes over a broad timescale. This thesis is dedicated to the development of novel structural MS approaches based on pulse covalent labelling and chemical cross-linking. Employing the developed quench-flow microfluidics apparatus, we performed footprinting experiments on proteins and protein complexes in timescale from a few microseconds to single seconds. Specifically, fast photochemical oxidation of proteins (FPOP) and novel fast fluoro alkylation of proteins (FFAP) techniques were utilized to track structural changes of myoglobin upon release of the prosthetic heme group....