63 research outputs found
RNAi phenotypes are influenced by the genetic background of the injected strain
No full textBackground: RNA interference (RNAi) is a powerful tool to study gene function in organisms that are not amenable to classical forward genetics. Hence, together with the ease of comprehensively identifying genes by new generation sequencing, RNAi is expanding the scope of animal species and questions that can be addressed in terms of gene function. In the case of genetic mutants, the genetic background of the strains used is known to influence the phenotype while this has not been described for RNAi experiments. Results: Here we show in the red flour beetle Tribolium castaneum that RNAi against Tc-importin alpha 1 leads to different phenotypes depending on the injected strain. We rule out off target effects and show that sequence divergence does not account for this difference. By quantitatively comparing phenotypes elicited by RNAi knockdown of four different genes we show that there is no general difference in RNAi sensitivity between these strains. Finally, we show that in case of Tc-importin a1 the difference depends on the maternal genotype. Conclusions: These results show that in RNAi experiments strain specific differences have to be considered and that a proper documentation of the injected strain is required. This is especially important for the increasing number of emerging model organisms that are being functionally investigated using RNAi. In addition, our work shows that RNAi is suitable to systematically identify the differences in the gene regulatory networks present in populations of the same species, which will allow novel insights into the evolution of animal diversity.DFG [FOR1234]Open-Access-Publikationsfonds 201
<i>foxQ2</i>has a key role in anterior head and central brain patterning in insects
No full textAnterior patterning of animals is based on a set of highly conserved transcription factors but the interactions within the protostome anterior gene regulatory network (aGRN) remain enigmatic. Here, we identify the foxQ2 ortholog of the red flour beetle Tribolium castaneum as novel upstream component of the aGRN. It is required for the development of the labrum and higher order brain structures, namely the central complex and the mushroom bodies. We reveal Tc-foxQ2 interactions by RNAi and heat shock-mediated misexpression. Surprisingly, Tc-foxQ2 and Tc-six3 mutually activate each other forming a novel regulatory module at the top of the aGRN. Comparisons of our results with those of sea urchins and cnidarians suggest that foxQ2 has acquired more upstream functions in the aGRN during protostome evolution. Our findings expand the knowledge on foxQ2 gene function to include essential roles in epidermal development and central brain patterning.</jats:p
Asymmetrically expressed axin required for anterior development in Tribolium
No full textCanonical Wnt signaling has been implicated in an AP axis polarizing mechanism in most animals, despite limited evidence from arthropods. In the long-germ insect, Drosophila, Wnt signaling is not required for global AP patterning, but in short-germ insects including Tribolium castaneum, loss of Wnt signaling affects development of segments in the growth zone but not those defined in the blastoderm. To determine the effects of ectopic Wnt signaling, we analyzed the expression and function of axin, which encodes a highly conserved negative regulator of the pathway. We found Tc-axin transcripts maternally localized to the anterior pole in freshly laid eggs. Expression spread toward the posterior pole during the early cleavage stages, becoming ubiquitous by the time the germ rudiment formed. Tc-axin RNAi produced progeny phenotypes that ranged from mildly affected embryos with cuticles displaying a graded loss of anterior structures, to defective embryos that condensed at the posterior pole in the absence of serosa. Altered expression domains of several blastodermal markers indicated anterior expansion of posterior fates. Analysis of other canonical Wnt pathway components and the expansion of Tc-caudal expression, a Wnt target, suggest that the effects of Tc-axin depletion are mediated through this pathway and that Wnt signaling must be inhibited for proper anterior development in Tribolium. These studies provide unique evidence that canonical Wnt signaling must be carefully regulated along the AP axis in an arthropod, and support an ancestral role for Wnt activity in defining AP polarity and patterning in metazoan development
Gr. 4 class 1, Beairsto School
No full textWayne Wutzki, Bill Spilchen, John Bidulka, George Ostashek, Alfred Kitzmann, Larry Joe, Alan McDougall, Bill Ingram, Barry Healey, Larry Seminuk, Ian Stead, Anne Lausman, Linda Brookbank, Judy Turanski, Jill Green, Arlene Anderson, Marnie Calvert, Elma Harvey, Jean Block Adriana Van Wyk, Joyce Woythe, Mary Knezwitch, Naomi Natsuhara, Betty Humphrey, Agnes Heilman, Vi Hubble, teacher, Sharon Desmarais, Shirley Watkins, Blanche Myette, Francis Dorchick, Sephie Pushkaunko, Norman Langton, Bill Costello, Peter Southward, Gordon Fox, Gary Wetherill, Ray Klein, Brian McIndoe, Michael Kidston
Hyperactive piggyBac transposase improves transformation efficiency in diverse insect species
No full textMorphogenesis and Genetic Regulation of the Insect Head
No full textNatural selection and the struggle for ecological niches were the driving force for the origin of a tremendous number of animal groups with different body shapes. The segmental organization of insects enhanced their evolutionary specialization. Adaptations led to different morphologies, for instance with respect to the head. The head, which carries the feeding apparatus and the main sensory centers enabling interaction with the environment, is one prerequisite for evolutionary success. The segmental structure of the insect head facilitated evolution of adaptations with respect to morphology. However, although the head is of great importance for the evolutionary success, there is no comprehensive understanding of the gene network regulating anterior head development until now. Furthermore, the genetic basis for the different insect head morphologies is unknown. Moreover, there are open questions concerning the segmental structure of the head and how morphogenetic movements lead to the adult head.
During the last years the red flour beetle Tribolium castaneum was developed as a major model organism for studying insect head development. Recent studies were able to successively uncover the genetic interactions of anterior head development. However, so far the gene set and its interaction are not comprehensively unraveled. Tc-foxq2 is a novel regulator of head development identified with a highly specific head phenotype in the iBeetle screen. Hence, I studied its potential function in the anterior head gene regulatory network. Another open question is the contribution of the different embryonic head regions to the adult head. To provide new insights regarding this question I wanted to provide new features for the Tribolium toolbox.
In this study I was able to show that Tc-foxq2 is specifically expressed in the anterior pre-segmental head region, similar to the conserved expression pattern, which is located at the anterior pole in metazoan species. Further, I was able to show that Tc-foxq2 is an upstream player within the anterior head gene regulatory network, forming a conserved patterning unit together with Tc-six3. Tc-foxq2 knock-down results in an increased cell death rate within the clypeo-labral region, which consequently leads to reduced labral buds at embryonic stages. This defect is also reflected in L1 larval cuticles showing a labrum that is strongly reduced or completely absent. Further, using neurogenic in vivo imaging reporter lines I was able to show that embryonic knock-down of Tc-foxq2 function leads to central body and mushroom body defects. Moreover, I was able to generate new reporter lines that drive expression of the chimeric H2Av::EGFP nuclear marker protein under control of the Tc-αTubulin1 promoter or the Tc-ribosomal protein subunit3 promoter. I analyzed the lines with respect to localization of the marker protein, signal distribution within the embryo, signal intensity in different developmental stages and tissues, and viability of the different transgenic lines. I could show that the lines are functional and that the Tc-αTubulin1 promoter line is best suited for being analyzed with light-sheet imaging. Finally, I was able to generate and to show functionality of new in vivo imaging lines for laser-induced cell marking and genetic cell marking
The Role of Substrate Temperature and Magnetic Filtering for DLC by Cathodic Arc Evaporation
No full textDiamond-like carbon (DLC) films were deposited using two different types of high current arc evaporation. The first process used a magnetic particle filter to remove droplets from the plasma. For the second process, the samples were put into a metallic cage which was placed directly above the plasma source. For both processes, we varied the substrate temperature from 21 to 350 °C in order to investigate the temperature effect. The samples were characterized using SEM, AFM, XPS, Raman Spectroscopy, Ellipsometry, Photometry, and Nano Indentation in order to compare both methods of deposition and provide a careful characterization of such DLC films. We found that the sp3 content and the hardness can be precisely adjusted by changing the substrate temperature. Furthermore, in the case of unfiltered deposition, the optical constants can be shifted in the direction of higher absorbance in order to produce black and hard carbon coatings
Production Efficiency Evaluation of Powder Coating Lines
No full textVõttes arvesse klientide kiireneva tarneaja soovi, on tarvis teha selle tarbeks mingeid tegevusi. Eelkõige on tarbekam vaadata ettevõtte tegevused üle ja teha hinnangud. Lähtuvalt ETS NORD AS ettevõttest, on otsustatud teha investeering tarneaja lühendamise suhtes. Selleks on uue automaatse värviliini ost, mis on olnud ka vajalikuks antud mure lahendamiseks. Tootlikkuse näitajad on suurenenud ning suudetakse värvida enam tooteid. Ettevõtte üheks heaolu faktoriks võib lugeda ehituse kulgu - kui on ehitus hoogsalt käimas ja ehitatakse pidevalt juurde, on ventilatsioon oluline tarbekaup, ning aitab ettevõttel paremini läbi lüüa. Antud juhul on ehituse trend aastate jooksul olnud tõusvas trendis, mis tõestab seda juhtumit. Autor on saanud selgemaks värviliinide tehnoloogia ning hinnanud olulisemaid tootmisefektiivsuse näitajaid. Saab väita, et automaatse liini puhul on tootlikkus näitajad suuremad kui manuaalse puhul. Selle tarbeks on mõõdetud ühe kui teise protsesside kestvusi ning tehtud järeldused. Seoses majanduslike aspektide seisukohast, on automaatsel liinil üldkulud suuremad, see-eest tööjõukulud ja riputamiseks mõeldud konksude maksuvus sarnane. Kvaliteedist lähtuvalt on mõlemal liinil ühtne järgimisviis pideva kontrolli tagamiseks ja selles erinevusi ei ole. Lõputöö eesmärgid on täidetud. Olemasoleva projektiga sai autor oskused olulisemate aspektide analüüsimisest, seadmete vaatlusest ja hinnangu tegemistest. Töö on olnud mõõdukalt mahukas ja on kindlasti kasulik tulevaste projektide läbiviimiseks.The following thesis is about Production Efficiency Evaluation of Powder Coating Lines. There is markable illustration that nowadays people attempt to get products as fast as possible. In addition, it have a solution - test a new method to eliminate those long waitings. ETS Nord AS is one of the biggest ventilation product and equipment producer and a seller in Estonia. The aim of this thesis is to investigate why a company invested to a new automatic based coating line. Also to use different measures to evaluate production relevant aspects, is the building trend somehow combined to show a company well-being. The important measures are: productivity, quality and flexibility. New coating line is compared to existing one - what has been already at working for 5 years. The thesis consist of 3 chapters. In the first one is described theoretical conception, which include description of a company, statistic of the construction build and info of a research object. The second chapter describes production efficiency aspects using a Park Son methods. The third one is the most complete and includes: technology of coating lines and hanging hooks, analysis of the technology and efficiency. Also the economical aspects - necessary costs and comparing the old and a new coating line as a production efficiency aspects. It can be said that a goal has been achieved through the measure procedures. The evaluations are done and there is a few summarizing aspects. A new coating line is more efficient than a manual operating line. Doing that thesis, author gained a lot of experience of evaluating, comparing and a observation. It definitely comes to be a favor of doing the following projects
A discussão do papel de uma teoria geral descritiva do direito na esteira do debate Hart-Dworkin
No full textO objetivo desta dissertação é discutir o papel de uma teoria geral descritiva do Direito nos moldes propostos por H.L.A. Hart na obra The Concept of Law. O trabalho parte das críticas de Ronald Dworkin ao Positivismo de Hart, passando pelas defesas elaboradas pelos autores positivistas e tendo como síntese a superação do debate Hart-Dworkin, com a conclusão de que as duas teorias ocupam espaços distintos na teoria jurídica. O primeiro capítulo é dedicado à reconstrução das críticas de Dworkin ao Positivismo Jurídico, expostas nas obras Taking Rights Seriously e Law’s Empire. Estas se dividem em duas frentes de ataque: a primeira consiste no argumento de que o Positivismo Jurídico vê o Direito como um sistema de regras, incapaz de dar conta da existência de princípios; a segunda defende que o Positivismo é incapaz de dar conta da existência de desacordos teóricos genuínos em Direito por estar comprometido com uma visão do Direito como simples questão de fato (plain fact view) e por estar preso ao aguilhão semântico (semantic sting). O segundo capítulo apresenta a defesa preliminar da teoria de Hart com base em John Gardner, Scott Shapiro e Joseph Raz. Neste capítulo, são desconstruídos os mitos que cercam o senso comum sobre o Positivismo, é elaborado um panorama geral da teoria positivista de Hart, bem como são distinguidas as duas principais correntes positivistas, quais sejam o inclusivismo e o exclusivismo. O terceiro e último capítulo é destinado à apresentação da resposta ao argumento de que o Positivismo é incapaz de explicar a existência de desacordos teóricos genuínos em Direito por estar comprometido com uma tese dos fatos sociais, demonstrando, com base em Luís Duarte d’Almeida e Kevin Toh, que essa crítica é fundada na compreensão equivocada distinção entre os pontos de vista externo e interno. Neste capítulo, também é analisado o aspecto normativo do Direito, com base na noção de fatos institucionais presente na obra Institutions of Law, de Neil MacCormick, autor que adota e leva adiante o projeto de uma sociologia descritiva proposto por Hart. O objetivo final do trabalho é demonstrar que não é necessário abandonar o Positivismo descritivo para dar conta das críticas de Dworkin, sendo que ainda há espaço na teoria jurídica para uma teoria geral descritiva do Direito nos moldes propostos por Hart.The aim of this work is to discuss the role of a general descriptive theory of law as proposed by H.L.A. Hart's The Concept of Law. The work begins with the critics of Ronald Dworkin to the positivism of Hart, going through the defenses prepared by positivist authors and having as its synthesis the overcoming of the Hart-Dworkin debate, concluding that the two theories occupy distinct areas in legal theory. The first chapter is devoted to the reconstruction of Dworkin's criticism of Legal Positivism, exposed in the works Taking Rights Seriously and Law's Empire. These fall into two attacking fronts: the first is the argument that Legal Positivism sees the law as a system of rules, unable to account for the existence of principles; the second argues that Positivism is unable to account for the existence of genuine theoretical disagreements in law for being committed to a plain fact view of the grounds of law and for being stung by the semantic sting. The second chapter presents the preliminary defense of Hart's theory based on John Gardner, Scott Shapiro and Joseph Raz. In this chapter, the myths surrounding the common sense of positivism are deconstructed, an overview of the positivist theory of Hart is elaborated, as well as the two main positivists strings are distinguished, namely inclusivism and exclusivism. The third and final chapter is for the submission of the response to the argument that Positivism is unable to explain the existence of genuine theoretical disagreements in law for being committed to a social fact thesis, demonstrating, based on Luís Duarte d'Almeida and Kevin Toh, that this criticism is based on the misapprehension of the distinction between external and internal points of view. In this chapter, it is also analyzed the normative aspect of law, based on the notion of institutional facts present in the work Institutions of Law, of Neil MacCormick, author who adopts and carries out the project of a descriptive sociology proposed by Hart. The final objective is to demonstrate that it not is necessary to leave descriptive positivism to account for Dworkin’s criticism, and to show that there is still room in legal theory to a descriptive general theory of law as the one proposed by Hart
RNAi phenotypes are influenced by the genetic background of the injected strain
No full textAbstract Background RNA interference (RNAi) is a powerful tool to study gene function in organisms that are not amenable to classical forward genetics. Hence, together with the ease of comprehensively identifying genes by new generation sequencing, RNAi is expanding the scope of animal species and questions that can be addressed in terms of gene function. In the case of genetic mutants, the genetic background of the strains used is known to influence the phenotype while this has not been described for RNAi experiments. Results Here we show in the red flour beetle Tribolium castaneum that RNAi against Tc-importin α1 leads to different phenotypes depending on the injected strain. We rule out off target effects and show that sequence divergence does not account for this difference. By quantitatively comparing phenotypes elicited by RNAi knockdown of four different genes we show that there is no general difference in RNAi sensitivity between these strains. Finally, we show that in case of Tc-importin α1 the difference depends on the maternal genotype. Conclusions These results show that in RNAi experiments strain specific differences have to be considered and that a proper documentation of the injected strain is required. This is especially important for the increasing number of emerging model organisms that are being functionally investigated using RNAi. In addition, our work shows that RNAi is suitable to systematically identify the differences in the gene regulatory networks present in populations of the same species, which will allow novel insights into the evolution of animal diversity.</p
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