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HajMohammadi, Sassan. "Development of FISH technology in pathological tissue." Thesis, University of Southampton, 1999. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.284578.
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Williams, Lisa. "Fluorescence in situ hybridisation (FISH) analysis of chromosomal aberrations in gastric tissue : the involvement of Helicobacter pylori." Thesis, Swansea University, 2004. https://cronfa.swan.ac.uk/Record/cronfa43067.
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Gastric cancer is a common cause of cancer death in the UK. It most often presents in patients when the disease is advanced, and hence treatment options are limited. As such, studies on the pre-malignant stages of gastric cancer, and interest in the mechanisms of the carcinogenic process (reactive oxygen species, ROS) and the agents that may drive the carcinogenic pathway {Helicobacter pylori infection), are important, with a view to improving disease outcome. This series of experiments has firstly shown, using CBMN assay +/- kinetochore staining and interphase FISH, that ROS causes aneuploidy of chromosomes 4, 8, 20 and 17(p53) in a human cell line. Secondly, gastric cells have been collected using endoscopic cytology brush techniques, and prepared, such that interphase FISH could be performed. Again, aneuploidy of chromosomes 4, 8, 20 and 17(p53) were detected in normal gastric mucosa, gastritis and intestinal metaplasia. The level of aneuploidy detected increased as disease severity increased. Amplification of chromosome 4, amplification of chromosome 20 and deletion of chromosome 17(p53) were the more significant findings. The degree of chromosomal abnormalities detected increased further, in a stepwise manner, when gastric dysplasia and gastric adenocarinoma cells were studied. Hence, a role for these abnormalities may exist in the initiation of, and the progression to, gastric cancer. The presence of H. pylori was also determined in the gastric tissue studied using histological analysis and PCR technology. Detection rates were comparable. The more virulent strain of H. pylori, Cag A, was found to be associated with increased disease pathology and chromosomal abnormalities, yet numbers were small. The amplification of chromosome 4 in gastric tissue was again highlighted in association with H pylori infection, hence it may reflect a role for chromosome 4 in the initiation of gastric cancer.
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Leversha, Margaret Anne. "Cytological estimations of molecular genetic difference : applications and implications of fluorescence in situ hybridisation mapping in the long arm of human chromosome 9." Thesis, University of Cambridge, 1994. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.337902.
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Stindl, Martin Maria Matthias. "Evaluation of ligation methods and the synthesis of a specific PNA-encoded peptide library." Thesis, University of Edinburgh, 2015. http://hdl.handle.net/1842/17927.
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Dysfunctional or over and under expressed enzymes play a crucial role in a variety of diseases. A tool that can identify dis-regulated enzymes in individual patients would be beneficial and would allow personalised treatment. For this purpose, a 10,000 membered ‘spit-and-mix’ PNA-encoded peptide library with a cell penetrating peptide was synthesised and interrogated with K562 cell lysate and intact K562 cells. This allowed the specific enzyme activity pattern for ABL tyrosine kinase from both inside a cell and a lysate to be obtained. Hybridisation of this library with a DNA-microarray resulted in bio-fouling by the cell lysate, thereby preventing analysis of the phosphorylation pattern. To allow extraction and purification of the peptide library from the cell lysates, a His-tag was incorporated into the library, and enabled successful library analysis. In addition to this 10,000 member library, a focused 100 PNA-encoded peptide library was synthesised. The library included peptide sequences known to be phosphorylated by specific tyrosine kinases deregulated in acute lymphoblastic leukaemia (ALL) with a PNA-tag complementary to a DNA microarray. Different ligation methods to conjugate the peptides to PNA-tags were screened – this included amide coupling, copper catalysed azide–alkyne cycloaddition, strain promoted azide–alkyne cycloaddition and Diels–Alder cycloaddition. The inverse electron demand Diels–Alder cycloaddition between a tetrazine and norbornene was chosen as the preferred ligation method, and the reaction conditions optimised. To purify the library from cell lysate, a His-tag was again coupled to each member using the strain promoted azide–alkyne cycloaddition. To test the tetrazine ligation, fluorescence in situ hybridisation (FISH) was used in cells, whereby a fluorophore was ligated onto a tetrazine–conjugated PNA probe. This was hybridised onto an mRNA in fixed cells. Results indicated that the ligation needed further optimisation.
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Somasekar, Amudha. "Fluorescence in situ hybridisation (FISH) analysis of chromosome 1 and gene expression levels of MAD2 and BUB1 levels in premalignant stages of gastric tissue." Thesis, Swansea University, 2011. https://cronfa.swan.ac.uk/Record/cronfa43141.
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Gastric cancer is a common cause of cancer death in the world. The mortality rate from gastric cancer is high in UK as it often presents late, often with local or distant metastasis. This makes the treatment options limited. The pathogenesis of gastric cancer occurs in a multi step pathway with pre cancerous conditions leading to cancer eventually. It is important to understand this carcinogenic process (aneuploidy and abnormal gene expression levels) and the driving forces (eg. Helicobacter pylori infection) which will enable us to alter the disease outcome. This series of experiments included cytogenetic investigation which involved obtaining gastric cells using brush cytology and using Fluorescent in situ hybridisation technique to look for aneuploidy levels of chromosome 1 and 4. These two chromosomes were chosen as chromosome 1 has been recently shown to be abnormal in early in premalignant stages of gastric cancer. Chromosome 4 was chosen as hyperploidy of chromosome 4 was the predominant chromosomal aberration in Barrett’s oesophagus. This study has shown that the aneuploidy level of chromosome 1 progressively increased with the progression of the histological stages according to the Correa’s premalignant gastric cancer pathway. Significant increase in aneuploidy levels of chromosome 1 was seen in H. Pylori associated gastritis, implying that H. Pylori play a very important role in the progression of the disease. Aneuploidy can occur due to various genetic defects that may potentially occur during mitosis. Spindle cell check points play a vital role in preventing the cells from proceeding to the anaphase stage if there is any defect in the kinetochore attachment. Certain genes like MAD2 and BUB1 are thought to be instrumental in controlling the spindle cell check points and it is believed a steady state of genes like MAD2 and BUB1 are required for this. In the second part of this study, the MAD2 and BUB1 expression levels were measured and correlated to the aneuploidy stages. There was no significant difference in their expression levels in patients with significant aneuploidy level. MAD2 levels were increased in H.Pyloriassociated gastritis, which implies that H. Pylori plays an important role in the pathogenesis of gastric cancer.
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Varela, Filipa Alexandra Pereira Rosa. "Tipificação de Mycoplasma mycoides subsp. mycoides SC e detecção da sua aderência a células epiteliais pulmonares." Master's thesis, Faculdade de Ciências e Tecnologia, 2010. http://hdl.handle.net/10362/5092.
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Dissertação apresentada para a obtenção do Grau de Mestre em Genética Molecular e Biomedicina, pela Universidade Nova de Lisboa, Faculdade de Ciências e TecnologiaA Peripneumonia Contagiosa Bovina (PPCB) é uma doença respiratória infecciosa, de grande relevo no âmbito da produção de gado bovino, causada pela bactéria Mycoplasma mycoides subsp. mycoides SC (MmmSC). Actualmente, a PPCB permanece endémica apenas em África, sendo aí responsável por perdas incontornáveis no sector pecuário. O último surto de PPCB detectado na Europa ocorreu em Portugal em 1999 mas, devido à sua natureza insidiosa, o risco de re-emergência é permanente. As estirpes de MmmSC associadas aos últimos surtos europeus de PPCB são tradicionalmente consideradas geneticamente muito homogéneas. No entanto, trabalhos recentes de tipificação molecular revelaram a existência de uma variabilidade genética intraspecífica superior ao que se esperava. A realização deste trabalho teve como base dois objectivos fundamentais: a tipificação de estirpes de MmmSC isoladas durante os últimos surtos de PPCB que ocorreram em Portugal entre 1993 e 1998, e a aplicação de um método alternativo e inovador para a detecção e quantificação de MmmSC, após a infecção de culturas celulares. Para a concretização do primeiro objectivo foi realizada uma análise da variabilidade polimórfica de três regiões VNTR (Variable Number of Tandem Repeats) existentes no genoma de estirpes de MmmSC. O locus VNTR4 comprovou ser o mais polimórfico, detectando-se quatro perfis distintos entre as estirpes portuguesas. O perfil VNTR4 do tipo “9” (numerado em função das repetições da sequência de consenso) revelou-se amplamente distribuído geograficamente e foi predominante entre os isolados. No entanto, observou-se uma segregação geográfica de perfis, dado que na região da Beira Litoral apenas foram encontradas estirpes com o perfil VNTR4 do tipo “8”. Estes resultados sugerem que podem ter ocorrido, pelo menos, dois eventos de re-emergência de PPCB em Portugal, entre 1993 e 1998. Para a realização do segundo objectivo deste trabalho foi optimizada uma técnica baseada na hibridação in situ com sondas de DNA fluorescentes (FISH) que permitiu visualizar micoplasmas aderidos a culturas celulares. Esta técnica, que nunca tinha sido usada previamente na detecção de MmmSC, comprovou ser adequada para futuras aplicações em estudos de citoaderência e para a detecção de micoplasmas em culturas celulares.
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Caburet, Sandrine. "Structure mosai͏̈que et instabilité de l'ADN ribosomal humain : implications dans la sénescence et la cancérogenèse." Paris 7, 2002. http://www.theses.fr/2002PA077038.
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Zheng, Yun-Ling. "Rapid prenatal diagnosis of common fetal aneuploidies by fluorescence in situ hybridisation." Thesis, University of Cambridge, 1993. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.318418.
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Birchall, Philip Simon. "Multicolour fluorescence in situ hybridisation to RNA in whole-mount Caenorhabditis elegans." Thesis, University of Cambridge, 1993. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.296668.
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Mohaddes, Ardebili Seyed Mojtaba. "Optimisation of interphase fluorescence in situ hybridisation for detection of common aneuploidies." Thesis, Connect to e-thesis, 1996. http://theses.gla.ac.uk/692/.
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Thesis (Ph.D.) - University of Glasgow, 1996.Ph.D. thesis submitted to the Faculty of Medicine, Department of Division of Developmental Medicine, University of Glasgow, 1996. Includes bibliographical references: p. 118-132. Print version also available.
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Hammond, David William. "Analysis of non-Hodgkin's lymphoma by conventional cytogenetics and fluorescence in-situ hybridisation." Thesis, University of Sheffield, 1995. http://etheses.whiterose.ac.uk/3064/.
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Cytogenetic analysis was performed on 40 non-Hodgkin's lymphoma (NHQ node biopsies. Chromosomes X, 3 and 12 were the most frequently gained; of the much rarer monosomies, loss of chromosome 13 was most common. Structural abnormalities primarily involved chromosomes 14,1,18,6 and 17. A markedly greater number of chromosome gains were associated with low-grade disease when compared to high-grade. In order to obtain further information from the cytogenetic analysis of the NHL karyotypes, the fluorescence in-situ hybridisation (FISH) technique was applied to the series. The activation state of additional X-chromosomes was examined and evidence that more than one X-chromosome was present in the active state in 4/9 cases was obtained. Further, in an apparent case of monosomy X, a marker was identified as an abnormal X-chromosome by chromosome painting. Interphase FISH was applied to NHL cells and numerical chromosome changes were identified; this approach was also attempted on aged bone marrow smears from acute lymphocytic leukaemia patients, in order to test the utility of the technique on archival material. Dual chromosome painting was used to elucidate the origins of add(14) chromosomes in 8 of the cases. In the control and two other cases the translocated material was demonstrated to be from chromosome 18, in two cases it was from chromosome 3 and in one case them was an insertion of chromosome 11 material. it was not possible to identify the origins of the translocated material in one NHL and in the final case the apparent add(14) was demonstrated not to contain chromosome 14 material. Structural abnormalities of chromosome 6 were investigated both by chromosome painting and by hybridisation of the MYB gene. The latter, which was initially mapped to 6q23 before hybridisation to NHL cells revealed previously unsuspected rearrangements. One case contained extrachromosomal chromatin bodies that appeared to be double minute chromosomes (dmin), which FISH analysis demonstrated to be derived from the X-chromosome and contain centromere-associated DNA. The significance of these results is discussed with reference to previously published series of NHL karyotypes.
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Sinclair, Paul Burdwood. "A fluorescence in-situ hybridisation and molecular investigation of leukaemia associated abnormalities of chromosome 6q." Thesis, University College London (University of London), 2002. http://discovery.ucl.ac.uk/1382604/.
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Deletions of the long arm of chromosome 6 are a recognised recurrent cytogenetic abnormality associated with ALL. Balanced rearrangements of 6q are less common but occur in leukaemias of both lymphoid and myeloid origin. With the aim of identifying genes that contribute to leukaemia through loss or rearrangement the breakpoints of translocations and deletions of 6q were mapped by FISH. Comparison between the mapped deletions led to the identification of a 4.8 Mb CDR and candidate tumour suppressor gene (GluR-6). Expression of GluR-6 was demonstrated to occur in haernatologic tissues and mutation analysis was performed on leukaemic samples with clonal deletions of the region. In one of 14 cases a base pair substitution that led to a change in amino acid sequence was found. The base pair substitution was also present in the patients' remission sample but was not seen in any of 20 other normal bone marrow samples analysed. Analysis of the translocations identified a variety of breakpoints between 6q15 and 6q27, with none positioned within the CDR. In seven cases breakpoints clustered within a 14 Mb region of 6q22-q23 but different subregions were defined for five analysed in detail. Complex rearrangements in one case of ALL appeared to result in translocation of one homologue of 6q and deletion of the second. Detailed FISH analysis defined a translocation breakpoint falling between two PACs that contained exons of a known tumour suppressor gene, the IGF2-R. Southern blot analysis failed to confirm disruption of the IGF2-R, but an IGF2-R-MRP8 fusion transcript was cloned by RACE PCR from the patients' c-DNA. Involvement of MRP8 in the translocation remains in doubt because attempts to confirm the presence of the fusion transcript were unsuccessful.
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Taylor, Clare Petronella Florence. "The use of fluorescence in situ hybridisation techniques in the diagnosis and prognosis of malignancy." Thesis, University College London (University of London), 1995. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.297282.
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Kim, Jeong-Soon. "Genomic analysis of sorghum by fluorescence in situ hybridization." Diss., Texas A&M University, 2003. http://hdl.handle.net/1969.1/1184.
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The reliability of genome analysis and proficiency of genetic manipulation in vivo and in vitro are increased by assignment of linkage groups to specific chromosomes, placement of centromeres, orientation with respect to telomeres, and linear alignment with respect to chromosomal features and dimensions. I undertook five studies aimed at integrating sorghum genomics and cytogenetics at several levels. The results help establish an entirely new "cyto-genomics" resource, impacts of which are likely to be broad. In the first study, I developed a FISH-based karyotyping system for Sorghum bicolor Moench. I used integrated structural genomic resources, including linkage maps and large-insert clonal libraries of sorghum genomic DNA to develop a 17-locus probe cocktail for simultaneous fluorescent in situ hybridization (FISH). This probe enabled facile identification of all chromosome pairs in mitotic chromosome spreads. Perhaps just as important, I established time-efficient means to select sorghum BAC clones for multi-probe FISH. Thus, an integrated cyto-genomics system for sorghum can be constructed without need of chromosome flow sorting or microdissection, both of which are difficult and costly. In the second study, hybridization of DNA clones from 37 different genomic regions enabled the assignment of linkage groups and orientation of linkage maps to chromosomes. Comparisons between genetic and physical distances throughout the genome enabled a new nomenclature for linkage group designation in sorghum. The results provide an integrated nomenclature system of Sorghum bicolor chromosomes and linkage groups. In the third study, I created high-resolution maps by FISH to pachytene bivalents for two linkage groups (B and H), and defined relationships between pericentromeric heterochromatin, centromeres, mapped markers and recombination rates. These relationships will help guide the development and use of sorghum genomics. In the fifth study, I used FISH in two ongoing gene-targeted efforts. For the maturity gene ma5 and fertility restoration gene rfl, I estimated physical lengths between currently available flanking molecular markers. This enables estimation of recombination densities in these regions and assessment of the applicability of map-based and -assisted cloning.
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Engelen, Johan Joseph Maria. "The application of micro-fish in clinical cytogenetics." Maastricht : Maastricht : Universiteit Maastricht ; University Library, Maastricht University [Host], 1998. http://arno.unimaas.nl/show.cgi?fid=6055.
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Szeto, Elaine. "Evaluation of fluorescence in situ hybridization (FISH) as a tool for screening of bladder cancer." Click to view the E-thesis via HKUTO, 2009. http://sunzi.lib.hku.hk/hkuto/record/B42904377.
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Prudent, Elsa. "Applications de l'hybridation in situ en fluorescence et stratégies moléculaires pour le diagnostic des infections bactériennes." Thesis, Aix-Marseille, 2018. http://www.theses.fr/2018AIXM0253/document.
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Une partie de ce travail de thèse a consisté à appliquer les méthodes de FISH pour l’étude de trois bactéries pathogènes intracellulaires. La viabilité de Bartonella henselae a été évaluée à partir de ganglions de patients atteints de la maladie des griffes du chat (CSD). Le faible taux d’ARN détecté par biologie moléculaire, la stérilité des cultures, l'absence de détection par analyses histologiques et FISH confirment que B. henselae n'est pas ou rarement viable dans les ganglions de patients atteints de CSD. Tropheryma whipplei, l’agent de la maladie de Whipple, a été identifié et localisé par FISH, dans les macrophages d’un ganglion et d’une biopsie pulmonaire, confirmant le diagnostic infectieux. Deux méthodes de FISH ont été testées pour détecter Coxiella burnetii dans des cas d’endocardites et d’infections vasculaires en utilisant des sondes oligonucléotidiques et des sondes PNA. Les résultats ont confirmé une meilleure efficacité des sondes PNA et démontré que les techniques de FISH sont plus sensibles que l’immunohistochimie pour le diagnostic des endocardites et des infections vasculaires à C. burnetii. Nous avons également évalué les stratégies moléculaires mises en place pour le diagnostic syndromique. Bien que la PCR conventionnelle à large spectre permette l'identification de micro-organismes fastidieux et anaérobies, la PCR spécifique en temps réel révèle une supériorité significative dans le diagnostic syndromique. En conclusion, ce travail a permis de démontrer l’efficacité et l’applicabilité de la FISH pour la détection bactérienne. Cette méthode peut être utilisée comme un outil complémentaire afin d'améliorer le diagnostic de microbiologie cliniqueWe applied FISH methods to the study of three intracellular pathogenic bacteria. The viability of Bartonella henselae was evaluated in a large series of lymph nodes from patients with cat scratch disease (CSD). The results obtained, associated with sterile cultures and negative histological analyzes and FISH, as well as the low level of RNA detected by molecular biology, provide evidence that B. henselae are not or are rarely viable in the lymph nodes of patients with CSD. Tropheryma whipplei has been identified by FISH in macrophages from one lymph node and for the first time in a pulmonary biopsy, confirming the diagnosis of infection. Two methods of FISH have been tested to detect Coxiella burnetii in cases of endocarditis and vascular infections using oligonucleotide and PNA probes. The results attested to the greater efficiency of PNA probes, and demonstrated that FISH were applicable for the diagnosis of C. burnetii endocarditis. We also evaluated the molecular strategies used for syndrome-driven diagnosis of infectious diseases. Although conventional broad-spectrum PCR allows for the identification of fastidious and anaerobic microorganisms, real-time specific PCR reveals a significant superiority in syndrome-driven diagnosis. The addition of specific PCRs in real time PCR would improve our molecular strategies, for example, in the case of the detection of Staphylococcus aureus for the diagnosis of lymphadenopathy. In conclusion, this work demonstrates the effectiveness and applicability of FISH for the identification of intracellular bacteria. This method can be used as an important complementary tool to the improvement of clinical microbiological diagnosis
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Salameh, Naser. "Physical mapping of maize chromosome 9 with the help of fluorescence in situ hybridization (FISH)." Beuren Stuttgart Grauer, 2005. http://d-nb.info/989891135/04.
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司徒柏沂 and Elaine Szeto. "Evaluation of fluorescence in situ hybridization (FISH) as a tool for screening of bladder cancer." Thesis, The University of Hong Kong (Pokfulam, Hong Kong), 2009. http://hub.hku.hk/bib/B42904377.
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Gordon, Alexander M. "Application of Fluorescence in situ Hybridization for Visualization and Quantification of Human Gastrointestinal Microbiota." Wright State University / OhioLINK, 2015. http://rave.ohiolink.edu/etdc/view?acc_num=wright1440846338.
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Zheng, Jianze. "Use of fluorescence in situ hybridization (FISH) for studying centromere organization and centric fusions in cattle." Title page, contents and abstract only, 1998. http://web4.library.adelaide.edu.au/theses/09A/09az63.pdf.
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Includes bibliograpical references (leaves 119-134). The most common chromosome abnormalities in live cattle are various Robertsonian translocations (centric fusions). Two hypotheses have been used to explain how monocentric Robersonian translocation chromosomes are generated: either direct formation, or evolution from dicentic chromosomes. Four main cattle procentric Satellite sequences were used as single and two-colour fluorescence in situ hybridization probes for studying the centromere organisation of cattle autosomes and the rearrangement in two cattle Robertsonian translocation chromosomes, the t(1:29) which is monocentric and found in numerous breeds, and the t(14:20) which is dicentric and found in 2 breeds.
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Chantot, Sandrine. "Aspects cytogenetiques des leucemies prolymphocytaires - b (lpl-b) : etude sequentielle en fish - multiplex (m - fish) des anomalies chromosomiques complexes dans un cas de lpl-b evolutive." Paris 5, 1999. http://www.theses.fr/1999PA05N086.
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Pedras, Maria Inês Machado. "Investigation of the regulation mechanisms for bioplastics production from industrial residues." Master's thesis, Faculdade de Ciências e Tecnologia, 2013. http://hdl.handle.net/10362/10863.
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Dissertação para obtenção do Grau de Mestre em
BiotecnologiaThe current high demand for plastics has become unsustainable. Polyhydroxyalkanoates are biopolymers stored by bacteria that can potentially replace modern plastics due to: wide range of applications; biodegradability; use of renewable resources as feedstock. High costs of current Polyhydroxyalkanoates production can be reduced using mixed cultures of organisms. Activated sludge from wastewater treatment plants is selected for Polyhydroxyalkanoates production through the imposition of cycles of intermittent feeding. In this study, the acclimation of activated sludge using synthetic volatile fatty acids (VFAs) as substrate resulted in a culture rich in Paracoccus spp. and unidentified filamentous bacteria. Low cost substrates such as sugarcane molasses (SM) or cheese whey (CW) can be employed as feedstock for further cost reduction. This requires an additional step before the microbial selection to ferment the feedstock into VFAs. In this work, the feedstock was changed from SM to CW. The population fed with SM was rich in Actinomycetaceae, while the population fed with CW was rich in Streptococcaceae, affecting the VFA composition. Consequently, the PHA-storing population and the polymer were affected. In the fermented SM (fSM) phase, the population was rich in Azoarcus (41.5 - 64.6%) and in the fCW phase the population was more diverse. Changing the pH in the fermentation reactor also affected the selection stage with an increase in Thauera and Azoarcus and a decrease in Paracoccus. A significant unidentified population of one layer sheet- forming bacteria was observed. Lastly, the occurrence of cell-to-cell communication (QS) in the selection stage was investigated. Possibly, QS molecules were detected when the carbon source was depleted. All steps of polyhydroxyalkanoate production are interconnected and for optimization, all stages must be studied and improved. Moreover, if QS proves to be involved in polyhydroxyalkanoate storage, the addition of QS molecules to the process may be explored for further optimization.
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Marzin, Youna. "Apport de l'hybridation in situ en fluorescence à l'étude des caryotypes complexes dans le myelome multiple." Brest, 2008. http://www.theses.fr/2008BRES3201.
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Le Myélome Multiple (MM) est une hémopathie maligne Caractérisée par une infiltration plasmocytaire de la moelle osseuse. Elle représente 10% des hémopathies malignes. Certaines anomalies chromosomiques sont des facteurs pronostiques utiles au suivi thérapeutique du patient. Cette étude a permis d’identifier et de préciser les anomalies chromosomiques de 94 patients atteints de MM entre Janvier 2000 et Décembre 2005, par des techniques conventionnelles, mais également par des techniques moléculaires de d’Hybridation In Situ en Fluorescence (FISH) et FISH 24 couleurs. Parmi ces anomalies, les réarrangements du chromosome 1 ont été mis en évidence chez 65 patients. La perte ip et le gain lq sont les anomalies les plus fréquentes, mais nous avons également mis en évidence des translocations sauteuses de ce chromosome chez 6 patients. Nous avons mis en évidence un réarrangement du gène IGH chez 39% de ces patients, certains réarrangements étant complexes. Par ailleurs, 56% des patients de la cohorte présentent une délétion 13q14, et 9,9% une délétion 17pl3. Enfin, les caryotypes de deux patients ayant rechuté en leucémie à plasmocytes ont été comparés. Une étude statistique portant sur les corrélations éventuelles entre les données cytogénétiques et les données cliniques et biologiques a révélé l’importance de la ploïdie. Certains patients présentent ainsi des caryotypes extrêmement complexes ne pouvant qu’aggraver la pathologie. Cette étude a permis de mettre en exergue des loci d’intérêt et propose des hypothèses de dérégulations géniques. Une meilleure connaissance des mécanismes d’évolution du MM pourrait ainsi permettre d’améliorer la prise en charge de ces patients pour lesquels il n’existe pas encore de thérapeutiques curativesMultiple myeloma (MM) is a malignancy cf the terminally-differentiated B cells that accounts for 10% of ail hematological malignancies. Some chromosomal abnormalities are important prognostic factors. This study is a cytogenetic work on 94 MM patients between January 2000 and December 2005, using conventional and molecular cytognetics technics, like Fluorescence In Situ Hybridization (FISH) and 24 colours FISH. Thus, 65 patients revealed chromosome 1 abnormalities. These abnormalities were most frequently a loss of lp arm and a gain of lq arm; jumping translocations were observed in 6 patients. Moreover, twenty-two patients (39%) had an 1Gb! translocation, which were sometimes complex, 56% had 13q14 deletion and 99% had l7pl3 deletion. Finally, we compared two MM karyotypes with their plasma cell leukemia karytopes. Statisticai study was used on cytogenetic, clinical and biological data and revealed the importance of ploidy. Some patients presented very complex karyotypes, probably increasing the pathology. This study showed loci of interest and proposed genetic deregulations hypotheses. Setter mechanism knowledge about MM evolution will permit to propose better treatments for this fatal pathology
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Sederberg, Maria C. "Physical Mapping of Ribosomal Genes in New World Members of the Genus Chenopodium Using Fluorescence in Situ Hybridization." BYU ScholarsArchive, 2008. https://scholarsarchive.byu.edu/etd/1629.
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The genus Chenopodium contains many economically important species in the New World, but is relatively understudied and poorly understood, especially in terms of evolutionary relationships. A better understanding of the structure of this genus could significantly help in breeding efforts on its cultivated members, notably the tetraploid C. quinoa and also certain varieties of C. berlandieri, also tetraploid. Of special concern is determining which diploid weed species are the most likely ancestors for C. quinoa, C. berlandieri, and the other tetraploid members of subsection Cellulata. The phylogeny can be understood in part by examining the ribosomal RNA loci and observing how many copies of the 5S and 45S loci each New World species contains. In this work, the 5S and 45S ribosomal RNA loci are characterized by means of fluorescence in situ hybridization in 23 Chenopodium species collected in the New World, with the 5S locus labeled red and the 45S locus labeled green. Based on these results, the pool of most likely candidate ancestor species for C. quinoa and C. berlandieri includes C. fremontii, C. incanum, C. neomexicanum, and C. watsonii.
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Tissir, Fadel. "Cartographie chez le rat :localisation régionale de gènes par hybridation in situ en fluorescence (FISH) et génération de nouveaux marqueurs génétiques." Doctoral thesis, Universite Libre de Bruxelles, 1998. http://hdl.handle.net/2013/ULB-DIPOT:oai:dipot.ulb.ac.be:2013/212064.
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Griffith, Brian Nelson. "The study of glucose-6-phosphate dehydrogenase (G6PD) gene regulation in HepG2 cells by glucose induction and the study of G6PD mRNA localization by fluorescent in situ hybridization (FISH)." Morgantown, W. Va. : [West Virginia University Libraries], 2002. http://etd.wvu.edu/templates/showETD.cfm?recnum=2511.
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Thesis (M.S.)--West Virginia University, 2002.Title from document title page. Document formatted into pages; contains viii, 100 p. : ill. (some col.). Vita. Includes abstract. Includes bibliographical references (p. 80-96).
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Berg, Johanna. "Visualisering av mikroorganismer i hårfolliklar från patienter med follikulit." Thesis, Umeå universitet, Biomedicinsk laboratorievetenskap, 2012. http://urn.kb.se/resolve?urn=urn:nbn:se:umu:diva-58521.
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Schmidt, Brian Friedrich. "The Optimization of the Catalyzed Reporter Deposition-Fluorescence in situ Hybridization (Card-Fish) Protocol for Future Use in Enumerating Populations of Cyanobacterial Picoplankton." Bowling Green State University / OhioLINK, 2010. http://rave.ohiolink.edu/etdc/view?acc_num=bgsu1274225695.
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Hultdin, Magnus. "Telomere analysis of normal and neoplastic hematopoietic cells : studies focusing on fluorescence in situ hybridization and flow cytometry." Doctoral thesis, Umeå University, Medical Biosciences, 2003. http://urn.kb.se/resolve?urn=urn:nbn:se:umu:diva-76.
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The telomeres are specialized structures at the end of the chromosomes composed of the repeated DNA sequence (TTAGGG)n and specific proteins bound to the DNA. The telomeres protect the chromosomes from degradation and end to end fusions. Due to the end-replication problem, the telomeric DNA shortens every cell division, forcing the cells into senescence at a critical telomere length. This process can be counteracted by activating a specialized enzyme, telomerase, which adds telomeric repeats to the chromosome ends leading to an extended or infinite cellular life span. Telomerase activity is absent in most somatic tissues but is found in germ cells, stem cells, activated lymphocytes and the vast majority of tumor cells and permanent cell lines. Hence, telomerase has been suggested as a target for cancer treatment as malignant cells almost exclusively express the enzyme and in that context telomere length measurements will be of great importance.
Telomere length is traditionally measured with a Southern blot based technique. A new method for telomere analysis of cells in suspension, called flow-FISH, was developed based on fluorescence in situ hybridization using a telomeric peptide nucleic acid (PNA) probe,
DNA staining with propidium iodide and quantification by flow cytometry. Flow-FISH had high reproducibility and the telomere length measurements showed good correlation with Southern blotting results. The flow-FISH technique also allows studies of cells in specific phases of the cell cycle and the replication timing of telomeric, centromeric and other repetitive sequences were analyzed in a number of cells. Like previous studies, centromeres were shown to replicate late in S phase while the telomere repeats were found to replicate early in S phase or concomitant with the bulk DNA, which is opposite to the patterns described in yeast.
In benign immunopurified lymphocytes from tonsils, high telomerase activity was found in germinal center (GC) B cells. This population also had high hTERT mRNA levels and displayed a telomere elongation as shown by flow-FISH and Southern blotting. Combined immunophenotyping and flow-FISH on unpurified tonsil cells confirmed the results.
Chronic lymphocytic leukemia (CLL), the most common leukemia in adults, can be divided into pre-GC CLL, characterized by unmutated immunoglobulin VH genes and worse prognosis, and post-GC CLL, with mutated VH genes and better prognosis. In 61 cases of CLL, telomere length was measured with Southern blotting and VH gene mutation status was analyzed. A new association was found between VH mutation status and telomere length, where cases with longer telomeres and mutated VH genes (post-GC CLL) had better prognosis
than CLL with short telomeres and unmutated VH genes (pre-GC CLL). A larger study of 112 CLL cases was performed using flow-FISH. The same correlation between telomere length and VH mutation status was found but gender seemed to be of importance as telomere length was a significant prognostic factor for the male CLL patients but not in the female group. Age of the patients and spread of disease seemed to affect the prognostic value of VH gene mutation status.
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Chang, Cindy Ma Schroeder Jane C. "Fluorescence in situ hybridization (FISH) and risk factors for non-Hodgkin lymphoma (NHL) subtypes defined by t(14;18) translocations and bcl-2 expression." Chapel Hill, N.C. : University of North Carolina at Chapel Hill, 2007. http://dc.lib.unc.edu/u?/etd,1212.
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Thesis (Ph. D.)--University of North Carolina at Chapel Hill, 2007.Title from electronic title page (viewed Mar. 26, 2008). "... in partial fulfillment of the requirements for the degree of Doctor of Philosophy in the Department of Epidemiology." Discipline: Epidemiology; Department/School: Public Health.
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Gaussen, Anthony. "Radiosensibilite predictive des tumeurs differenciees de la thyroide : etude comparative par le fish et la clonogenicite cellulaire : correlation avec la sensibilite des lymphocytes." Paris 11, 1998. http://www.theses.fr/1998PA11T040.
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Traboulsi, Abdel-Meneem. "Étude à moyen-débit de la localisation d'ARNm dans les cellules humaines." Thesis, Montpellier, 2017. http://www.theses.fr/2017MONTT117.
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La localisation d’ARNm a été découverte en 1983 dans les ovocytes et les embryons des ascidies. Depuis, plusieurs exemples d'ARN localisés ont été trouvés dans de nombreux organismes, y compris les plantes, les levures, les champignons, les insectes, les poissons et les mammifères. Les ARNm localisés contribuent à de nombreuses fonctions biologiques, telles que le développement embryonnaire, la division cellulaire asymétrique, la migration cellulaire, la signalisation, la plasticité neuronale et plein d’autres ...Jusqu'à présent, quelques études ont analysé la localisation d’ARNm de manière systématique. Trois d'entre eux ont été effectués chez la drosophile pendant l'embryogenèse, l'oogenèse ou le stade larvaire et ont analysé environ 16000 ARN au total. Les deux autres études ont été réalisées dans des cellules de mammifères et ont analysé près de 1000 ARNm chacune. Ces études ont montré l'importance de la localisation d’ARNm dans les cellules humaines et son implication dans différents processus biologiques. L'objectif de ma thèse était donc d'augmenter le débit des techniques FISH à l’échelle de molécule unique (smFISH) et d'étudier la localisation d’ARNm dans les cellules HeLa de manière systématique.Une limitation de smFISH est le coût de sondes fluorescentes, qui limite le nombre d'ARNm qui peut être analysé. Par conséquent, j'ai développé un protocole alternatif dans lequel des sondes pour de nombreux gènes ont été synthétisées comme un pool d'oligonucléotides (40 par gène en moyenne, plus de 12000 au total). Les sondes spécifiques d’un ARNm donné ont ensuite été amplifiées par PCR et converties en simple brin par transcription in vitro. J'ai généré un protocole complet, à partir de la conception de la sonde et jusqu'à l'acquisition de l'image. Je me suis intéressé à l’étude des ARNm du cycle cellulaire. En effet, les gènes du cycle cellulaire ont été largement étudiés au niveau de la protéine, mais on sait peu de choses sur la localisation de leurs ARNm. Pendant la mitose, les cellules subissent d'importantes modifications morphologiques et la traduction locale pourrait être un moyen d'atteindre la localisation des protéines. Le screening sur ces ARNm est en cours.Parallèlement à ces expériences, j'ai réalisé des expériences de smFISH sur 100 gènes choisis au hasard et 50 régulateurs de la transition G2/M du cycle cellulaire, en utilisant un protocole de smFISH classique. Dans cette configuration, on disposait d'une collection de lignées cellulaires HeLa, dans laquelle chaque cellule contient un chromosome artificiel bactérien avec le gène d'intérêt marqué au GFP. Par conséquent, en utilisant des sondes qui s'hybridaient à la séquence GFP, je pourrais utiliser le même ensemble de sondes marquées pour étudier la localisation de tous les ARNm. Un autre avantage est que la localisation des protéines pourrait être évaluée simultanément. Mes résultats indiquent que 4 ARNm ont montré une localisation spécifique lors du screening de 100 gènes choisis d’une manière aléatoire et 15 ARNm parmi les 54 régulateurs de la transition G2 / M. Ces ARNm appartiennent à cinq classes de localisation: "blobs", qui sont des agrégats d'ARNm cytoplasmiques; «clusters», qui sont des zones de concentration locale élevée d'ARNm, mais où une molécule unique d’ARNm peut encore être résolu; «nuclear membrane », où les ARNm se concentrent autour de l'enveloppe nucléaire; "spindle", qui sont des ARNm accumulés sur l'appareil de division mitotique, “spots" qui sont des agrégats d'ARNm cytoplasmiques où une molécule unique d’ARNm ne peut pas être résolu, et qui sont plus grands que les blobs. La colocalisation entre l'ARNm et la GFP, qui suggère une traduction locale, n'a été trouvée que pour 1 ARNm.Ces screenings aléatoires et ciblés effectués à petite échelle montrent une fréquence et une diversité inattendues dans les modèles de localisation d’ARNm. Cela ouvre la voie pour effectuer des screenings à plus grande échelleMRNA localization was discovered in 1983 in ascidian oocytes and early embryos. Since then many examples of localized RNAs have been found in many organisms, including plants, yeast, fungi, insects, fish and mammals. Localized mRNAs contribute to many biological functions, such as embryonic patterning, asymmetric cell division, cell migration, signaling, neuronal plasticity and others…Until now, only few studies analyzed RNA localization in a systematic manner. Three of them were done in Drosophila, during embryogenesis, oogenesis or larval stage and analyzed around 16000 mRNAs in total. The two other studies were done in mammalian cells and analyzed nearly 1000 mRNAs each. These studies opened a door and raised questions regarding the importance of mRNA localization in human cells and its implication in different biological processes. The goal of my thesis was thus to increase the throughput of single molecule FISH techniques (smFISH) and to study mRNA localization in HeLa cells in a systematic manner.One limitation in smFISH is the cost of the fluorescent oligonucleotide probes, which limits the number of mRNAs that can be analyzed. Therefore, I developed an alternative protocol in which probes for many genes were synthesized as a pool of oligonucleotides (40 per gene in average, more than 12000 in total). Gene-specific probes were then amplified by PCR and converted into single strand by in vitro transcription. I generated a complete protocol, starting from probe design and up to image acquisition. I was interested in studying cell cycle genes. Indeed, cell cycle genes have been extensively studied at the protein level but little is known concerning the localization of their mRNAs. During mitosis, cells go through important morphological modifications and local translation could be a mean of achieving protein localization. This screen is ongoing.In parallel to these experiments, I performed a smFISH based screen on 100 randomly chosen genes and 50 regulators of the G2/M transition of the cell cycle, using a traditional smFISH protocol. In this set-up, I took advantage of a library of HeLa cell lines, in which each cell line contains a bacterial artificial chromosome with the gene of interest tagged with GFP. Therefore, using oligonucleotides hybridizing to the GFP sequence, I could use the same probe set to study the localization of all the tagged mRNAs. A further advantage is that protein localization could be assessed simultaneously. My results indicate that two mRNAs showed a specific localization when screening 100 random genes, and 16 mRNAs among the 50 regulators of the G2/M transition. These mRNAs belong to five localization classes: "blobs", which are cytoplasmic mRNA aggregates; "clusters", which are areas of high local mRNA concentration but where individual mRNA can still be resolved; "nuclear envelope", where mRNAs concentrate around the nuclear envelope; "spindle", which are mRNAs accumulating on the cell division apparatus during mitosis, “spots" which are cytoplasmic mRNA aggregates where individual mRNA can’t be resolved and are bigger than blobs. Interestingly, colocalization between mRNA and GFP, which suggests local translation, was only found for 1 mRNA.These random and targeted screens performed at small-scale show an unexpected frequency and diversity in mRNA localization patterns, therefore pointing to new functions related to this process. This will stimulate future studies aiming at performing screenings at a higher scale
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Doaré-Lebrun, Elodie. "Caractérisation de la microflore des raisins par méthodes FISH et PCR-TTGE : Application à la résolution des goûts terreux dans les vins." Compiègne, 2005. http://www.theses.fr/2005COMP1619.
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Depuis quelques années, des déviations aromatiques qualifiées de terreuses ou moisies sont observées dans les vins de plusieurs régions viticoles françaises, en Beaujolais, dans le Val de Loire ou dans le Bordelais. Tous les types de vins peuvent être touchés. La principale molécule responsable de ces altérations est la géosmine. Elle est vraisemblablement produite très précocement à la vigne, par des micro-organismes présents sur les raisins. Par ailleurs, divers micro-organismes sont connus pour produire cette molécule, comme des bactéries actinomycètes, appartenant notamment au genre Streptomyces ou des moisissures du genre Penicillium. Nous avons mis au point des outils méthodologiques pour la détection simple, rapide et précoce des micro-organismes responsables de la production des molécules à l'origine des défauts terreux dans les vins. La technique FISH est utilisée pour la détection précoce des Streptomyces sur les raisins grâce à la sonde HGC. Cependant, aucun Streptomyces n'est détecté par la méthode FISH sur nos échantillons de raisins, bien qu'ils soient vraisemblablement présents, puisqu'ils ont été détectés par méthodes de microbiologie classique. La présence de spores de moisissures très majoritaires masque vraisemblablement la présence de ces micro-organismes. Concernant les moisissures, la détection des espèces productrices de géosmine n'est pas possible par la technique FISH par manque de sondes spécifiquesSince a few years, sensorial defects defined as earthy or musty odours have been detected in some wines of several French wine-producting regions like Beaujolais, Loire Valley and Bordeaux. All types of wines can be concerned. The molecule responsible for these defects is geosmin. It is produced very early in the vineyeards by micro-organisms present on the grapes. We also know that actinomycete bacteria, including the Streptomyces genus or fungi belonging to the Penicillium genus cans produce geosmin. We have developed methodological tools for an easy, quick, and early detection of the micro-organisms responsible for the production of the molecules causing the earthy defects in wines. The FISH method is used for the early detection of Streptomyces on grapes using the HGC probe. However, no Streptomyces has been detected with the FISH technique on our grapes'samples, even if they were probably there as they have been detected by classic mircrobiological methods. This Gan be caused by a large majority of fungal spores in the samples, masking the presence of these micro-organisms. Regarding the moulds, the detection of geosmin-producing species is not possible du to the lack of specific probes
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Funari, Mariana Ferreira de Assis. "Identificação de deleções do gene SHOX: comparação das técnicas de FISH, análise de microssatélites e MLPA." Universidade de São Paulo, 2009. http://www.teses.usp.br/teses/disponiveis/5/5135/tde-09022010-174156/.
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O gene SHOX (short stature homeobox containing gene), expresso em altos níveis nas células osteogênicas, é fundamental para o desenvolvimento ósseo e para a determinação da altura. Haploinsuficiência do SHOX é responsável por vários fenótipos que envolvem a baixa estatura, como a síndrome de Turner, a discondrosteose de Léri-Weill e a baixa estatura idiopática. Cerca de dois terços das haploinsuficiências são causados por deleções. Neste trabalho, foi realizada uma comparação entre três técnicas para detecção de deleções do SHOX: a hibridação in situ com fluorescência (FISH), o estudo de microssatélites e o multiplex ligationdependent probe amplification (MLPA). Nos pacientes sem deleção do SHOX, foi realizado um rastreamento para identificação de mutações de ponto no gene que levassem à sua haploinsuficiência. Foram analisados seis pacientes com discondrosteose de Léri-Weill (DLW) e 20 com baixa estatura desproporcionada (BED). Na técnica de FISH, os cromossomos metafásicos obtidos a partir de cultura de linfócitos foram hibridados com o cosmídio LLNOYCO3M34F5. DNA genômico extraído a partir de leucócitos de sangue periférico foi submetido à análise de microssatélites e MLPA. Foram amplificados seis marcadores de microssatélites (repetições CA, DYS290, DXYS10093, DXYS10096, DXYS233 e DXYS234) e o MLPA foi realizado de acordo com as instruções dos kits SALSA MLPA P018-C1 e P018-D1 SHOX. Estes kits contêm oito sondas específicas para o gene SHOX e 13 para a área do SHOX, localizada a jusante do gene. O seqüenciamento direto da região codificadora do gene foi realizado nos pacientes sem deleção. Todos os pacientes com DLW apresentaram deleções envolvendo todo o gene. Entre os pacientes com BED, apenas um (5,0%) apresentou uma deleção intragênica envolvendo os exons 4, 5 e 6a. Os resultados das três metodologias foram concordantes na maioria dos casos, exceto em dois casos. No primeiro caso, inicialmente o FISH não identificou uma deleção envolvendo todos os éxons em um paciente com DLW. No segundo, uma deleção envolvendo os exons 4, 5 e 6a, identificada em uma paciente com BED, foi detectada apenas pelo MLPA. Ainda entre os pacientes com BED, três (15%) apresentaram deleção da região do marcador DXYS10096, a 3 do gene. Outros três (15%) pacientes apresentaram mutações de ponto identificadas pelo seqüenciamento direto: a mutação p.Tyr35X, que resulta na substituição de uma tirosina por um códon de parada prematuro; a p.Arg147His localizada na região do homeodomínio e a NM_000451:c.1236 -10T>C que se encontra a 10 nucleotídeos antes do início do éxon 5. Em uma comparação das três metodologias, o FISH foi considerado a técnica mais trabalhosa e com menor sensibilidade, levando até oito dias para sua realização. A análise por microssatélites requer o estudo dos progenitores, além de um grande número de marcadores para a análise de regiões extensas. O MLPA detectou todas as deleções, sendo considerada a metodologia mais sensível. Ele apresentou também menor custo e tempo de execução, além de possibilitar a estimativa do tamanho da deleção. Desta forma, o MLPA foi considerado a melhor metodologia para investigação inicial dos pacientes com DLW e BED.The SHOX gene (short stature homeobox containing gene), expressed at high levels in osteogenic cells, is essential for bone development and growth process. SHOX haploinsufficiency is responsible for several phenotypes involving short stature, such as Turner syndrome, Léri-Weill dyschondrosteosis (LWD) and idiopathic short stature. Deletions are responsible for 2/3 of SHOX haploinsufficiency. In this study, a comparison among three techniques for detection of SHOX deletions: fluorescence in situ hybridization (FISH), microsatellites analysis and multiplex ligationdependent probe amplification (MLPA) was performed. A screening for point mutations that could lead to haploinsufficiency was performed in patients without SHOX deletion. Six patients with Léri-Weill dyschondrosteosis (LWD) and 20 with disproportionate short stature (DSS) were analyzed. FISH analysis was performed using the cosmid LLNOYCO3\"M\"34F5 and metaphase spreads obtained from lymphocytes culture. Genomic DNA extracted from peripheral blood leukocytes was used to microsatellite and MLPA analysis. Six microsatellite markers (CA repeats, DYS290, DXYS10093, DXYS10096, DXYS233 and DXYS234) were amplified by PCR and MLPA was performed according to the manufacturers instructions for SALSA MLPA P018 and P018-C1-D1 SHOX kits. These kits contain 8 specific probes for SHOX gene and 13 for \"SHOX area, which is located downstream of the gene. The direct sequencing of entire encoding region was performed in patients with no SHOX deletions. All patients with LWD presented deletions involving the entire gene. One (5.0%) patient with DSS, presented an intragenic deletion involving exons 4, 5 and 6a. The results of the three methods were concordant in most cases, except in two cases. In the first case, a patient with DLW, the FISH did not identify a deletion involving all SHOX exons. In the second case, a deletion of exons 4, 5 and 6a in a patient with BED was identified only by MLPA. Other 3 (15%) DSS patients had deletion in SHOX area, in the DXYS10096 marker. Other three (15%) patients presented a point mutation identified by direct sequencing: p.Tyr35X, which replaces a tyrosine for a premature stop codon, p.Arg147His located in the homeodomain region and NM_000451: c.1236-10T> C which is 10 nucleotides before the exon 5. In a comparison of three methods, the FISH technique was considered the more laborious and less sensitive, taking until eight days to obtain the results. The microsatellite analysis requires the parents DNA study. In addition, several markers are essential for the analysis of extensive regions. The MLPA was considered the most sensitive methodology since it detected all deletions. It also presented lower cost and execution time, and allowed the estimation of the size of the deletion. Thus, the MLPA was considered the best approach for initial investigation of LWD and DSS patients.
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Julian, Katia. "Caractérisation moléculaire d'inversions péri- et paracentriques et analyse de leurs effets sur la méiose d'individus porteurs hétérozygotes." Toulouse 3, 2010. http://thesesups.ups-tlse.fr/1286/.
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Les inversions sont des anomalies chromosomiques de structure résultant d'une double cassure sur un même chromosome et d'un retournement à 180° du segment chromosomique ainsi généré. Ces inversions sont dites péricentriques si les points de cassure sont localisés de part et d'autre du centromère, ou paracentriques dans le cas contraire. Chez l'Homme, la fréquence estimée des inversions péricentriques est de 0. 012% à 0. 07% et de 0,01% à 0,05% pour les inversions paracentriques (McKinlay-Gardner et Sutherland, 2004). Les porteurs hétérozygotes ne présentent généralement pas d'altération phénotypique mais sont susceptibles de rencontrer des problèmes de reproduction en raison de perturbations du processus méiotique conduisant à des troubles de la spermatogenèse et/ou à la production de gamètes génétiquement déséquilibrés. De nombreuses inversions ont été identifiées chez l'Homme et dans les espèces animales d'élevage. Cependant, peu d'études de ségrégation méiotique d'inversions ont été publiées à ce jour chez l'Homme (par exemple Morel et al. 2007), et aucune dans les espèces animales d'élevage. L'analyse méiotique de différentes inversions identifiées chez le porc dans le cadre du programme national de contrôle chromosomique mis en œuvre par notre laboratoire présente un double intérêt. Le premier est zootechnique. En effet, lors de la mise en évidence d'une inversion chez un futur reproducteur, sa réforme était jusqu'à présent systématiquement conseillée aux éleveurs. Toutefois, l'application de ce principe peut s'avérer difficile et/ou non optimal car elle peut conduire à l'élimination d'individus par ailleurs génétiquement intéressants et ainsi induire une baisse de l'efficacité des programmes de sélection. Raisonner l'utilisation des reproducteurs sur la base d'une prédiction de l'effet potentiel des remaniements nous semble une alternative raisonnable. L'un des objectifs de la Thèse est donc la mise au point et l'évaluation d'une méthode de prédiction basée sur l'estimation du pourcentage de gamètes déséquilibrés dans des échantillons de semence. Le second objectif est cognitif. En effet, des difficultés d'ordre technique et/ou éthique rendent difficile l'acquisition de certaines connaissances concernant le comportement méiotique de remaniements chromosomiques chez l'Homme. C'est le cas par exemple de l'effet du sexe du parent porteur sur le comportement et les produits de la méiose en présence d'une inversion. Le recours à un modèle animal (le porc en l'occurrence, dont la structure chromosomique est proche de celle de l'Homme) permet de s'affranchir de certaines contraintes, et d'apporter de nouvelles connaissances dans des domaines non encore documentés. Un autre objectif de la Thèse est, dans cet esprit, de réaliser une étude comparée des ségrégations méiotiques " mâle " et " femelle " d'une même inversion et ainsi de déterminer l'influence du sexe du porteur sur les effets de l'anomalie. L'analyse des profils de ségrégation méiotique " mâle " a été réalisée par la méthode de SpermFISH (hybridation in situ en fluorescence sur noyaux de spermatozoïdes décondensés). Les ségrégations méiotiques femelles ont pour leur part été étudiées par analyse de métaphases II d'ovocytes de truies maturés in vitro. Dans les deux cas, des sondes moléculaires de type BAC ont été utilisées. Les études d'appariement méiotique ont été réalisées à partir de biopsies testiculaires réalisées chez un verrat porteur d'une inversion péricentrique du chromosome 4. Ces échantillons ont été analysés par des méthodes d'immunocytologie à l'aide d'anticorps permettant de détecter spécifiquement certaines protéines. Les études de ségrégation méiotique de 7 inversions de tailles et de types différents ont été réalisées : les pourcentages de gamètes génétiquement déséquilibrés variaient de 0,6% à 4%, suggérant que ces inversions ont un impact très limité sur la reproduction. De manière surprenante, la proportion de gamètes anormaux produits n'est pas corrélée avec la taille du fragment inversé, comme cela est le cas chez l'Homme. La comparaison des profils de ségrégation " mâle " et " femelle " pour une inversion du chromosome 4 n'a pas non plus montré de différence de production de gamètes déséquilibrés entre les deux genres (4% et 3,6% respectivement). Le sexe du porteur ne semble donc pas être un facteur déterminant pour la production de gamètes anormaux. L'analyse des phases précoces de la méiose pour un animal porteur de la même inversion péricentrique du chromosome 4 à montré un comportement méiotique particulier du bivalent: plus de 90% des cellules montraient en effet un appariement non homologue de la région inversée. De plus, des évènements de crossing over dans la région inversée du chromosome 4 n'ont été observés que dans 5% des cas. Le comportement des chromosomes est responsable de la faible recombinaison dans la région inversée et donc de la faible proportion de gamètes génétiquement déséquilibrés estimée (4% en moyenne). Ces résultats demandent néanmoins à être confirmés par d'autres analyses. L'étude en cours des phases précoces de la méiose chez deux porteurs d'inversions péricentriques du chromosome 8 devraient nous permettre très prochainement d'approfondir nos connaissances sur le comportement méiotique des chromosomes en présence d'inversion. Par ailleurs, la mise au point de nouvelles techniques au sein du laboratoire, telles que la production de cellules souches induites (iPS) porcines, nous permet aussi d'envisager à moyen terme l'étude de l'intégralité du processus méiotique mâle en présence d'une inversionInversions are structural chromosomal rearrangements formed when a chromosome breaks in two places and the fragments reverses orientation. The inversion is pericentric if the breaks occur on either side of the centromere, paracentric in other cases. Prevalence estimates in human population range from 0. 012% to 0. 07% for pericentrics and from 0,01% to 0,05% for paracentrics (McKinlay-Gardner et Sutherland, 2004). Usually, both para- and pericentric inversions are phenotypically harmless, but the presence of an inversion can occasionally lead to severe reproductive disorders due to spermatogenesis impairements and the production of genetically unbalanced gametes. Many cases of inversions have been described in humans as well as in other animal species. However, relatively few studies have reported meiotic segregation pattern analyses of inversions in males (for example Morel et al. 2007), and none were reported on domestic animals. Meiotic analyses of various inversions identified in the pig species, thanks to the national program of chromosomic control, have a double interest. The first is zootechnical. When an inversion is detected in a boar, the breeders are always advised to sacrifice the animal. However, it can be difficult to eliminate individuals with a high genetic value and then induce a lowering in the efficiency of selection programms. Using a prediction of the potential effects of rearrangements to advise the breeders seem to be a good alternative. One of the thesis purpose is then to create a predictive method based on the percentage of unbalanced gametes in semen samples. The second interest is more linked to basic research. Indeed, it can be difficult to acquire knowledge on meiotic behaviour of chromosomal rearrangements in Man due to technical and/or ethical reasons. For example, little is known on an " sex " effect on meitoic behaviour during meiosis for inversion carriers. The use of an animal model (like the pig species) is a great opportunity to analyze the meiotic behaviour of chromosomes in inversion carriers, and then to have a better understanding on not well documented domains. That is why an other purpose of this thesis was to carried a comparative study of meotic segregations in males and females carriers of the same anomaly, and then determine a potential " sex " effect. Meotic segregation pattern analyses were carried out using the SpermFISH technique (fluorescent in situ hybridization on decondensed sperm nuclei) for males, and FISH on metaphase II oocytes matured in vitro for females. In both cases, molecular probes (Bacterial Artificial Chromosome) were used. Studies of pairing behaviour were carried out on testicular samples of an inversion of chromosome 4 carrier, using immunocytological techniques (use of specific antibodies against proteins of interest). Meotic segregation pattern analyses were carried out for 7 inversions (different length of the inverted fragment and different type) : percentage of unbalanced gametes ranged from 0,6% to 4%, suggesting a limited impact on reproduction. Surprisingly, the length of the inverted fragment is not correlated to the proportion of unbalanced gametes (unlike in Man). Comparison of " male " and " female " segregation profiles for an inversion of chromosome 4 did not show either any difference concerning the production of unbalanced gametes (4% and 3,6% respectively). The gender of the carriers do not seem to be a major factor for the production of unbalanced gametes. Analyses of early stages of meiosis for a carrier of the same pericentric inversion showed a specific meiotic behaviour of the bivalent : a non homologous pairing of the inverted region was seen in more than 90% of the cells analysed. Moreover, crossing overs in the inverted region were relatively rare (5 % of the cells). This chromosomal behaviour explains the low recombination rate in the inverted region, and then the low proportion of unbalanced gametes produced (mean of 4%). These results have to be confirmed by other analyses. Studies of the early stages of meiosis in two carriers of pericentric inversions of chromosome 8 should allow in the future a better understanding of the meiotic behaviour for inversion carriers. Moreover, the use of new techniques, such as production of induced pluripotent stem cells (iPS), could allow, in the future, analyses of the whole male meiotic process in inversion carriers
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Lemos, Samira Salomão. "Identificação e quantificação de bactérias em dentes decíduos com necrose pulpar por meio da técnica da Hibridização in Situ Fluorescente (FISH)." Universidade Federal de Juiz de Fora, 2014. https://repositorio.ufjf.br/jspui/handle/ufjf/1352.
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CAPES - Coordenação de Aperfeiçoamento de Pessoal de Nível Superior
Diferentes métodos de identificação têm mostrado que a comunidade microbiana em infecções endodônticas apresenta uma grande diversidade. No entanto, o conhecimento dos perfis microbianos de infecções endodônticas em dentes decíduos ainda é limitado. Portanto, este estudo teve como objetivos avaliar qualitativa e quantitativamente a presença de micro-organismos orais em amostras de canais radiculares dentes decíduos com necrose pulpar, e verificar a correlação entre eles e sua associação com sinais e sintomas clínicos e radiográficos. Um total de 31 crianças, de quatro a dez anos (idade média = 6,29 ± 1,27 anos), foi incluído neste estudo. A presença de 12 patógenos selecionados (Actinobacillus actinomycetemcomitans, Campylobacter rectus, Enterococcus faecalis, Fusobacterium nucleatum, Porphyromonas gingivalis, Prevotella intermedia, Prevotella nigrescens, Streptococcus, Streptococcus mutans, Streptococcus sobrinus, Tannerella forsythia e Treponema denticola) em 31 canais radiculares de 31 dentes com necrose pulpar foi avaliada por meio da técnica de hibridização in situ fluorescente (FISH), que identificou e quantificou os micro-organismos. Medidas descritivas foram usadas para descrever os dados relativos à densidade (cél/mL X 108) obtida para cada micro-organismo testado. O teste de correlação de Pearson avaliou a correlação entre os micro-organismos identificados e o teste t de Student verificou a associação entre os sinais e sintomas clínicos e radiográficos e os micro-organismos detectados. Adotou-se um nível de significância de 5% (p < 0,05). Todos os patógenos testados foram identificados em todas as amostras. O somatório das densidades médias totais de todas as espécies bacterianas testadas e do gênero Streptococcus representou 80,57% da comunidade microbiana total. Os resultados do teste t de Student demonstraram que houve uma diferença significante (p = 0,02) entre no número médio das densidades observadas na espécie T. denticola no grupo com presença de dor e o grupo com ausência de dor. Também foram observadas diferenças significantes no número médio das densidades da espécie P. nigrescens e presença de edema e P. nigrescens e ausência de edema; assim como para o gênero Streptococcus e presença de edema e Streptococcus e ausência de edema (p = 0,04; p = 0,04, respectivamente). A técnica de FISH confirmou a característica polimicrobiana da infecção endodôntica em dentes decíduos com a presença de bactérias anaeróbicas obrigatórias e anaeróbicas facultativas e do gênero estreptococos.
Different microbial identification methods have shown that the microbial community in endodontic infections presents a great diversity. However, the knowledge of the microbial profiles of endodontic infections in primary teeth is still limited. Therefore, the aim of this study was to evaluate, qualitative and quantitatively, the presence of oral microorganisms in samples from root canals primary teeth, to assess the correlations among them, and to determine their association with clinical and radiographic signs and symptoms. A total of 31 children, 4 to 10 years old (mean age = 6.29 ± 1.27 years old), was involved in this study. The presence of 12 selected pathogens (Actinobacillus actinomycetemcomitans, Campylobacter rectus, Enterococcus faecalis, Fusobacterium nucleatum, Porphyromonas gingivalis, Prevotella intermedia, Prevotella nigrescens, Streptococcus, Streptococcus mutans, Streptococcus sobrinus, Tannerella forsythia, and Treponema denticola) in 31 infected root canals from 31 primary teeth with pulp necrosis was studied by using the fluorescent in situ hybridization (FISH) technique which identified and quantified the microorganisms. Descriptive measures were used to present the data related to the density (cel/mL X 108) for each microorganism tested. The Pearson correlation test assessed the correlation among the microorganisms identified and the Student t test assessed the association between the clinical and radiographic signs and symptoms and the pathogens detected. The significance level was set at 5% (p < 0.05). All the tested pathogens were detected in all samples. The total sum of the mean densities of all the bacteria species and Streptococcus genus represented 80.57% of the entire microbial community. The results of Student's t test showed that there was a significant difference (p = 0.02) between the average number of densities observed in the species T. denticola in the group with pain and the group with no pain. Significant differences were also observed in the average number densities of the species P. nigrescens and presence of edema and P. nigrescens and no edema; and for Streptococcus and genus Streptococcus and edema and no edema (p = 0.04, p = 0.04, respectively). The FISH technique confirmed the polymicrobial characteristic of the endodontic infection in primary teeth with the presence of obligate and facultative anaerobic bacteria and of Streptococcus genus.
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Faria, Karina de Cassia [UNESP]. "Análise citogenética comparativa em espécies de morcegos dos gêneros Molossus (Molossidae), Artibeus, Platyrrhinus, Sturnira, Glossophaga, Phyllostomus e Carollia (Phyllostamide) - Chiroptera (Mammalia)." Universidade Estadual Paulista (UNESP), 2003. http://hdl.handle.net/11449/92536.
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faria_kc_me_sjrp.pdf: 1587104 bytes, checksum: 5b515af867e10b2a892710a2b48a4662 (MD5)Para verificar a ocorrência de homologias cromossômicas entre espécies de Molossidae e Phyllostomidae (Chiroptera), foram realizadas análises comparativas do bandamento G e, hibridização in situ fluorescente genômica e telomérica nas espécies Carollia perspicillata (Carolliinae), Glossophaga soricina (Glossophaginae), P. hastatus (Phyllostominae), Sturnira lilium, Platyrrhinus lineatus e Artibeus planirostris (Stenodermatinae) de Phyllostomidae e, Molossus ater e Molossus molossus (Molossinae) de Molossidae. As análises do bandamento G evidenciaram que, à exceção de C. perspicillata, todas as demais espécies de Phyllostomidae compartilham homologias cromossômicas com as espécies de Molossidae. As espécies A. planirostris, P. lineatus e S. lilium apresentam 29 dos 32 braços cromossômicos de Molossus. Estas espécies compartilham dois cromossomos inteiros e os braços de oito cromossomos meta ou submetacêntricos destas espécies de Stenodermatinae apresentam homologias com cromossomos acrocêntricos e subtelocêntricos de Molossus. G. soricina também apresenta 29 braços cromossômicos de Molossus, sendo que três cromossomos inteiros são compartilhados e os braços de sete cromossomos de G. soricina apresentam homologias com cromossomos acrocêntricos e subtelocêntricos de Molossus. Todos os braços cromossômicos de P. hastatus apresentaram homologias com os cromossomos de Molossus. P. hastatus e Molossus compartilham cinco cromossomos inteiros e os braços de oito cromossomos de P. hastatus apresentam homologias com os cromossomos acrocêntricos e subtelocêntricos de Molossus. Estes resultados parecem sugerir uma proximidade maior das espécies de Molossidae com P. hastatus (Phyllostominae). Além de rearranjos do tipo fusões e inversões pericêntricas, outros rearranjos cromossômicos não determinados justificam as diferenças entre os cariótipos das espécies de...
To verify the occurrence of chromosome homologies between species of Molossidae and Phyllostomidae (Chiroptera), comparative analysis of the G-banding and, genomic and telomeric fluorescence in situ hibridization were accomplished in the species Carollia perspicillata (Carolliinae), Glossophaga soricina (Glossophaginae), Phyllostomus hastatus (Phyllostominae), Sturnira lilium, Platyrrhinus lineatus and Artibeus planirostris (Stenodermatinae) of Phyllostomidae and, Molossus ater and Molossus molossus (Molossinae) of Molossidae. The analysis of the G-banding evidenced that except C. perspicillata all the other species of Phyllostomidae share chromosome homologies with the species of Molossidae. The species A. planirostris, P. lineatus and S. lilium present 29 of the 32 chromosome arms of Molossus. These species share two whole chromosomes and the arms of eight meta or submetacentric chromosomes of these species of Stenodermatinae present homologies with acrocentric and subtelocentric chromosomes of Molossus. G. soricina also presents 29 chromosome arms of Molossus, in a way that three whole chromosomes are shared and the arms of seven chromosomes of G. soricina present homologies with acrocentric and subtelocentric chromosomes of Molossus. All of the chromosome arms of P. hastatus presented homologies with the chromosomes of Molossus. P. hastatus and Molossus share five whole chromosomes and the arms of eight chromosomes of P. hastatus present homologies with the acrocentrics and subtelocentrics chromosomes of Molossus. These results seem to suggest a larger proximity of the species of Molossidae with P. hastatus (Phyllostominae). Besides rearrangements like fusions and pericentric inversions, other not determined chomosome rearrangements justify the differences between the karyotype of the compared species of Phyllostomidae and Molossidae. The comparative genomic hybridizations with Molossus...(Complete abstract click electronic access below)
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Milhiet, Elodie. "Nanospectroscopie de molécules d’intérêt biologique." Paris 11, 2007. http://www.theses.fr/2007PA112150.
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La spectroscopie de molécule unique joue aujourd’hui un rôle majeur dans de nombreux domaines allant de la physique fondamentale à la biologie. Dans ce contexte, mes travaux ont conduit au développement théorique et instrumental de deux méthodes d’investigation orientées vers la biologie. La première visait à caractériser la dynamique de complexation du calcium par la sonde calcique fluorescente Oregon Green Bapta5N communément employée pour l’analyse des signaux intracellulaires. Pour y parvenir, nous avons développé un dispositif expérimental de spectroscopie de corrélation de fluorescence à un et deux photons présentant une sensibilité proche de la molécule unique. Grace à ce dernier, nous avons pu étudier plusieurs aspects de la photophysique de la sonde et avons évalué ses limites ainsi que l’intérêt de l’appliquer in vivo. La seconde a consisté à développer une technique d’Hybridation In-Situ de Fluorescence (FISH) semi-quantitative afin de cartographier l’expression de gènes dans le cerveau de drosophiles adultes. Nous avons surmonté deux difficultés majeures, en obtenant, pour la première fois, des résultats reproductibles et semi-quantitatifs chez la drosophile adulte. Je présente ici une nouvelle approche où l’amplification enzymatique a été remplacée par une détection optimisée et un protocole réduisant l’impact de l’autofluorescence. Des résultats sur divers gènes exprimés dans le cerveau des drosophiles adultes y sont exposés au même titre qu’une étude visant à mieux comprendre une pathologie de retard mental. Pour conclure, j’ai mis en évidence la capacité de notre technique à résoudre des sondes uniques ce qui ouvre la voie vers de nouvelles applicationsSingle-molecule-like spectroscopy plays a major role in many domains, from fundamental physics to biology. In this framework, my dissertation focuses on instrumental and theoretical developments of two biological-related applications. The first experiment aims at characterizing the dynamics of calcium binding by the fluorescent calcium probe Oregon-green Bapta5N commonly employed in cell signaling analysis. To achieve it, I have developed an experimental set-up of fluorescence correlation spectroscopy that exhibits sensitivity close to that of single-molecule detection. Either monophotonic or biphotonic excitations can be used. I have investigated the several aspects of the photophysics of the probe and evaluated the interest and limitations of such an approach for future in-vivo measurements. The second one is devoted to the development of a semi-quantitative Fluorescent In-Situ Hybridization (FISH) technique for mapping gene expression in the adult drosophila brain. Two difficulties have to be solved. First, we succeeded in obtaining reproducible results with drosophila adult brain. Secondly, while most of the FISH protocols are not quantitative since they need a strong enzymatic, we achieved semi-quantitative detection of RNA probes. I will present results on a new approach for which enzymatic detection is replaced by a sensitive detection and a protocol which reduces autofluorescence contribution. Results will be presented for several genes in adult drosophila brain to validate the methods as well as an interesting application on a mental retardation disease. To conclude, I show that the method exhibits a single RNA sensitivity which opens the way to new applications
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Brocardo, Graciela Aparecida. "Avaliação do comprimento dos telômeros em células infectadas pelo vírus HTLV-I utilizando a técnica hibridização in situ fluorescente e citometria de fluxo (Flow-FISH)." Universidade de São Paulo, 2008. http://www.teses.usp.br/teses/disponiveis/5/5167/tde-25032009-174020/.
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INTRODUÇÃO: A Leucemia/Linfoma de células T do adulto (ATL) é uma doença linfoproliferativa crônica com transformação clonal predominantemente de linfócitos TCD4+, causada pelo vírus linfotrópico T humano do tipo I (HTLV-I). A ATL se desenvolve em 3-5% dos portadores do vírus HTLV-I, após longo período de latência clínica, acompanhado de expansão clonal dos linfócitos infectados. As células da ATL apresentam várias anormalidades cromossômicas, semelhantes àquelas resultantes de disfunção telomérica e a instabilidade genômica contribui para o desenvolvimento da ATL. Para entender o papel do encurtamento telomérico na oncogênese da ATL, avaliamos o comprimento dos telômeros de linfócitos TCD4 e TCD8 em portadores do vírus HTLV-I e em portadores de ATL. RESULTADOS: Não foi evidenciada diferença significativa no comprimento de telômero dos subtipos linfocitários TCD4+ e TCD8+ entre portadores do vírus HTLV-I e indivíduos saudáveis, assim como, entre portadores de ATL e indivíduos saudáveis. Entretanto, quando incluímos na análise a variável idade, evidenciamos redução significativa do comprimento do telômero com a idade em portadores do vírus HTLV-I e maior perda telomérica nos portadores do vírus HTLV-I e portadores de ATL em relação aos indivíduos saudáveis de mesma idade, embora a diferença entre os grupos não atinja o nível de significância estatística. Estes resultados podem ser explicados pelo fato de que as células dos indivíduos infectados pelo vírus HTLV-I apresentam maior taxa proliferativa devido à ação viral, mesmo em estado de latência clínica. A perda telomérica em função da idade nos portadores de ATL não demostrou-se significativa devido ao pequeno número de casos analisados em decorrência da raridade da doença. Entretanto, quando analisamos o comprimento telomérico nos subtipos linfocitários de portadores de ATL, evidenciamos acentuada perda telomérica na célula maligna e valores próximos ao limite superior esperado para a idade no subtipo linfocitário não transformado, demonstrando que a disfunção telomérica deve estar associada à transformação celular. Estabelecemos valores de referência de comprimento telomérico dos subtipos linfocitários TCD4+ e TCD8+ de indivíduos saudáveis, definidos por faixa etária. CONCLUSÃO: Nossos resultados demonstram que portadores do vírus HTLV-I apresentam maior perda telomérica em função da idade que indivíduos saudáveis, mas, sem refletir significância estatística e clínica. Entretanto, portadores de ATL apresentam perda acentuada de comprimento de telômero na célula maligna, demonstrando que a determinação do comprimento de telômero pode auxiliar futuramente o monitoramento dos indivíduos infectados pelo HTLV-I, indicando conversão à doençaINTRODUCTION: Adult T-cell Leukemia/Lymphoma (ATL) is a chronic lymphproliferative disease with clonal transformation predominantly of the TCD4+ lymphocytes, caused by the Human T lymphotropic virus type-I (HTLV-I). ATL develops itself in 3-5% of HTLV-I carriers after a long period of clinical latency accompanied by clonal expansion of the infected lymphocytes. The ATL cells present several chromosomic abnormalities, similar to those resulting from telomere dysfunction and the genomic instability contributes to the development of ATL. In order to understanding the role of telomeric shortening in the ATL oncogenesis, we assessed the length of telomeres of lymphocytes TCD4 and TCD8 in HTLV-I carriers and in ATL carriers. RESULTS: No significant difference was evidentiated in the telomere length of lymphocytary subtypes TCD4+ and TCD8+ between HTLV-I carriers and healthy subjects, as well as, between ATL carriers and healthy subjects. However, when the age variable was included in the analysis, we observed significant decrease of telomeric length with age progression in HTLV-I carriers and higher telomeric loss in HTLV-I carriers and ATL carriers when compared to healthy subjects of the same age, although the difference between groups does not reach the level of statistic relevance. These results may be explained by the fact that the cells of HTLV-I infected subjects present higher proliferative rate due to the viral action, even during clinical latency. Age-related telomeric loss in ATL carriers did not manifest itself as significant due to the small number of analyzed cases as a consequence of the diseases rareness. However, when the telomere length on the lymphocytary subtypes of ATL carriers was analyzed, we evidentiated accentuated telomeric loss in the malignant cell and values close to the age-expected upper limit in the nontransformed lymphocytary subtype, demonstrating that the telomere dysfunction may be associated to the cellular transformation. We have determined reference values of telomere length for lymphocytary subtypes TCD4+ and TCD8+ on healthy subjects, defined by age range. CONCLUSION: Our results demonstrate that HTLV-I carriers present higher telomeric loss due to age than healthy subjects, however, with no reflection in clinical and statistical significance. Nevertheless, ATL carriers present accentuated loss of telomere length in the malignant cell, demonstrating that the telomere length determination may, in the future, assist in the monitoring of HTLV-I infected subjects, indicating conversion to the disease
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Krämer, Dorothee Charlotte Agathe. "Investigation of Mammalian Chromatin Folding at Different Genomic Length Scales using High Resolution Imaging." Doctoral thesis, Humboldt-Universität zu Berlin, 2019. http://dx.doi.org/10.18452/19929.
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Chromatin ist ein Makromolekül, dessen Genregulation innerhalb des räumlich eingeschränkten Zellkerns organisiert werden muss. Die Genomorganisation ist eng mit Genaktivierung und Genrepression verknüpft. In den vergangenen Jahren wurde gezeigt, dass die DNA hierarchisch organisiert ist. Die Faltung läuft in aufeinander folgenden Schritten ab, wobei jede Organisationsebene sowohl zur räumlichen Komprimierung, als auch zur Genregulation beiträgt. In dieser Dissertation wurden mit Hilfe von hochauflösender Mikroskopie verschiedene Ebenen der 3D Chromatinorganisation auf Einzelzell-Basis untersucht. Auf der kleinsten Organisationsebene wurde die Struktur zweier, nebeneinander liegender topologischer Domänen (TADs) am Sox9-Lokus erforscht. Mit Hilfe von Fluoreszenz in situ Hybridisierung (FISH) in 3D Zellen, sowie Cryoschnitten in embryonalen Stammzellen von Mäusen konnten Interaktionen zwischen den benachbarten TADs festgestellt werden. FISH in Zellen mit genomischen Duplikationen, zeigte das Entstehen von zwei unterschiedlichen, durch die Duplikation entstandenen, Konformationen. Unter Verwendung von FISH wurden long-range Kontakte, die zuvor mit GAM entdeckt wurden, untersucht und es zeigte sich, dass sie häufig zwischen TADs die regulatorischen Domänen enthalten auftreten. Zudem zeigte sich die Bildung von Clustern zwischen mehreren, weit auseinander liegenden, regulatorischen Elementen. Dies lässt unter Umständen auf das Entstehen von regulatorischen Zentren zwischen diesen Enhancer-reichen Regionen schließen. Weitere Untersuchungen zeigten Veränderung der sogenannten Super-Enhancer Cluster in unterschiedlichen Zelltypen. Des Weiteren sind Super-Enhancer TADs sehr dekondensiert und wurden häufig an Splicing-Speckle Regionen vorgefunden.Chromatin needs to organize gene regulation whilst fitting into the confined space of the nucleus. Chromatin organization is therefore intertwined with gene activation and silencing. In recent years many advances in the field of chromatin architecture have been made showing that chromatin is organized hierarchically. Folding occurs in subsequent units, where each level of organization contributes to the spatial compaction of DNA and gene regulation. In this dissertation different levels of 3D chromatin organization were analysed using single-cell, high-resolution imaging. On the smallest scale, the 3D organization of two neighbouring Topologically Associating Domains (TADs) at the Sox9 locus was investigated. Performing Fluorescence in situ Hybridization (FISH) in 3D and cryosectioned mouse embryonic stem cells, extensive contacts between the two neighbouring TADs across the TAD boundary were detected. Applying FISH in a cell line bearing a genomic duplication within the Sox9 locus, the occurrence of two different conformations that result from the duplication was shown. Recent evidence from GAM showed the formation of long-range, multimer contacts between distal regulatory elements. Investigating the occurrence of long-range contacts between super-enhancer TADs in single cells by FISH, showed that they establish frequent interactions at close spatial distances. Furthermore the formation of clusters containing distal super-enhancer TADs could be demonstrated, indicating the possibility of higher-order regulatory hubs between these enhancer-rich regions. Further investigation showed that super-enhancer regions form different clusters in different cell types. Finally, it was shown that super-enhancers are highly decondensed and preferentially located at splicing speckles.
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Latuf, Juliana de Paulo 1985. "Investigação da frequencia de núcleos 45,X por meio de hibridização in situ com fluorescência (FISH) em linfócitos e mucosa oral de homens normais e sua aplicação a mosaicos 45,X/46,XY." [s.n.], 2013. http://repositorio.unicamp.br/jspui/handle/REPOSIP/308157.
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Orientadores: Andréa Trevas Maciel-Guerra, Vera Lúcia Gil da Silva LopesDissertação (mestrado) - Universidade Estadual de Campinas, Faculdade de Ciências Médicas
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Resumo: Quadros de ambiguidade genital e esterilidade com cariótipo 46,XY podem ser devidos a mosaico com linhagem 45,X não detectável no cariótipo em linfócitos de sangue periférico. Quando essa linhagem não é detectada, esses indivíduos deixam de ser investigados em relação a uma série de problemas clínicos. Este trabalho teve como objetivo verificar se a hibridação in situ com fluorescência (FISH) em células de mucosa oral poderia ser empregada para detectar criptomosaicismo com linhagem 45,X em indivíduos com cariótipo 46,XY. A casuística foi composta por 19 homens saudáveis com idades entre 20 e 30 anos e cinco pacientes com distúrbios da diferenciação do sexo (DDS) com idades entre 5 e 23 anos, quatro com mosaico 45,X/46,XY e um com disgenesia testicular 46,XY associada a déficit de crescimento. Após confirmar que os jovens saudáveis tinham cariótipo 46,XY em 50 metáfases de linfócitos de sangue periférico, foi realizada análise por FISH com sondas específicas para os cromossomos X e Y em 1.000 núcleos interfásicos de linfócitos de sangue periférico e 1.000 de mucosa oral, seguida de comparação da proporção de núcleos contendo apenas o sinal do cromossomo X nos dois tecidos. A mesma análise foi feita nos cinco pacientes com DDS. A distribuição da proporção dos núcleos interfásicos de linfócitos e mucosa oral contendo apenas o sinal do X nos jovens saudáveis foi compatível com a distribuição normal, e número superior a 12:1.000 em linfócitos e 13:1.000 em mucosa oral devem ser considerados indicativos de mosaicismo em nosso laboratório. A frequência desses núcleos nos dois tecidos não diferiu significativamente (p=0,6855). Nos cinco pacientes com DDS a frequência de núcleos contendo apenas o sinal do X diferiu significativamente da observada em indivíduos normais em linfócitos (p=0,0008) e mucosa oral (p=0,0008). No paciente com cariótipo prévio 46,XY a linhagem 45,X foi confirmada por FISH em metáfases, e em um dos casos de mosaicismo foram detectadas linhagens celulares adicionais. Também não houve diferença significativa entre a frequência de núcleos contendo apenas o sinal do X nos dois tecidos desses pacientes (p=0,3750). Estes resultados indicam que a pesquisa de mosaicismo com linhagem 45,X em indivíduos com DDS ou esterilidade e cariótipo 46,XY pode ser feita por meio de FISH em mucosa oral, com vantagens evidentes em termos de custo e rapidez, além de ser feita a partir de tecido obtido de modo não invasivo
Abstract: Ambiguous genitalia and sterility with a 46,XY karyotype may be due to mosaicism with a 45, X karyotype not detectable in peripheral blood lymphocytes. When this cell line is not detected, these individuals fail to be investigated over a range of clinical problems. This study aimed to verify whether fluorescence in situ hybridization (FISH) in cells from buccal smear could be employed to detect cryptomosaicism with a 45,X cell line in individuals with a 46,XY karyotype. The sample consisted of 19 healthy men aged 20 to 30 years and five patients with disorders of sex development (DSD) aged 5 to 23 years, four with mosaicism 45,X/46,XY and one with testicular dysgenesis 46, XY associated with growth deficiency. After confirming that the healthy young men had a 46,XY karyotype in 50 metaphases from peripheral blood lymphocytes, FISH analysis with probes specific for chromosomes X and Y was done in 1,000 nuclei from peripheral blood lymphocytes and 1,000 from buccal smear, followed by comparison of the proportion of nuclei containing only the signal of the X chromosome in these tissues. The same analysis was performed in five patients with DDS. The distribution of the proportion of interphase nuclei of lymphocytes and buccal smear containing only the X signal in healthy young was consistent with normal distribution; a number greater than 12:1,000 in lymphocytes and 13:1,000 in buccal smear should be considered indicative of mosaicism in our laboratory. The frequency of these nuclei in both tissues did not differ significantly (p = 0.6855). In patients with DDS the frequency of nuclei containing only the X signal differed significantly from that observed in normal individuals both in lymphocytes (p = 0.0008) and buccal smear (p = 0.0008). In the patient with a prior 46,XY karyotype, a 45,X cell line was confirmed by FISH in metaphases, and in one case of mosaicism additional cell lines were detected. There was also no significant difference between the frequency of nuclei containing only the X signal in the two tissues of these patients (p = 0.3750). These results indicate that investigation of mosaicism with 45,X cell line in individuals with 46,XY DSD or sterility can be done by FISH in cells from buccal smear, with obvious advantages in terms of cost and speed, using a tissue obtained noninvasively
Mestrado
Ciencias Biomedicas
Mestra em Ciências Médicas
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Spiluttini, Béatrice. "Interaction du snARN U1 de l'épissage avec l'ARN polymérase II." Phd thesis, Université Pierre et Marie Curie - Paris VI, 2009. http://tel.archives-ouvertes.fr/tel-00814598.
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Les ARNs non codants sont des régulateurs de l'expression génétique à plusieurs niveaux. Chez la bactérie et chez la souris, des ARNs non codants (6S et B2) ont la propriété de se lier à l'ARN polymérase et d'inhiber son activité. Afin de déterminer si l'ARN polymérase II (RNAPII) humaine était associée à des ARNs non codants, une immunoprécipitation anti-RNAPII a été réalisée sur des cellules HeLa mitotiques. Les ARNs co-immunoprécipités ont été purifiés et marqués et l'ARN U1 s'est trouvé particulièrement enrichi par rapport au contrôle. Cette co-immunoprécipitation reflète l'association de la snRNP U1 avec la RNAPII. Pour vérifier cette association sur un site de transcription actif, des lignées ont été établies avec l'insertion en multiples copies d'un gène à un site unique, créant ainsi un unique super site de transcription visualisable par FISH (Fluorescence In Situ Hybridization). Deux lignées distinctes ont été créées, l'une avec un gène comportant un intron, l'autre avec le même gène où l'intron comporte trois mutations ponctuelles abolissant l'épissage. Alors que les snARNs U2, U4, U5 et U6 sont absents du site non épissé, l'ARN U1 est enrichi de la même façon indépendamment de l'épissage. La présence des protéines spécifiques de la snRNP U1 indique que la snRNP U1 est recrutée au complet au site de transcription. Ces résultats laissent supposer un rôle pour l'association RNAPII - U1snRNP dans l'épissage cotranscriptionnel.
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Calvó, Perxas Laia. "A study on the phylogeny and the ecology of ammonia-oxidizing bacteria using a new molecular marker based on the gene amoB." Doctoral thesis, Universitat de Girona, 2005. http://hdl.handle.net/10803/7866.
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L'agricultura i la industrialització han causat un augment significatiu del nombre d'ambients rics en amoni. La presència de compostos nitrogenats redueix la qualitat de l'aigua, causant problemes de toxicitat, deteriorant el medi ambient i fins i tot afectant la salut humana. En conseqüència, la nitrificació s'ha convertit en un procés global que afecta al cicle del nitrogen a la biosfera. Els bacteris oxidadors d'amoni (AOB) són els responsables de l'oxidació de l'amoni a nitrit, i juguen un paper essencial en el cicle del nitrogen.Els primers oxidadors d'amoni foren aïllats a finals del segle XIX, però la lentitud del seu creixement i les dificultats per cultivar-los feren que fins als anys 80, amb els primers estudis emprant el gen 16SrDNA, no s'assolís un coneixement complert d'aquest grup bacterià. Actualment les bases de dades contenen multitud d'entrades amb seqüències corresponents a AOB.
L'objectiu d'aquest treball era trobar, desenvolupar i avaluar eines útils i fiables per a l'estudi dels AOB en mostres ambientals.
En aquest treball primer descrivim la utilització de la hibridació in situ amb fluorescència (FISH), mitjançant l'aplicació de sondes amb diana en el 16SrRNA dels AOB. La FISH ens va permetre detectar i recomptar aquest grup bacterià; no obstant, aquest mètode no permetia la detecció de noves seqüències, pel que es necessitava una nova eina.
Amb aquesta intenció vam aplicar la seqüència de la sonda Nso1225 en una PCR. El fet d'amplificar específicament un fragment del 16SrDNA dels AOB va suposar el desenvolupament d'una nova eina molecular que permetia detectar la presència i diversitat d'aquests bacteris en ambients naturals. Malgrat tot, algunes seqüències pertanyents a bacteris no oxidadors d'amoni del subgrup β dels proteobacteris, eren també obtingudes amb aquesta tècnica. Així mateix, un dels inconvenients de l'ús del 16SrDNA com a marcador és la impossibilitat de detectar simultàniament els AOB que pertanyen als subgrups β i γ dels proteobacteris.
El gen amoA, que codifica per la subunitat A de l'enzim amoni monooxigenasa (AMO), era aleshores àmpliament utilitzat com a marcador per a la detecció dels AOB. En aquest treball també descrivim la utilització d'aquest marcador en mostres procedents d'un reactor SBR. Aquest marcador ens va permetre identificar seqüències de AOB en la mostra, però la necessitat de detectar amoA mitjançant clonatge fa que l'ús d'aquest marcador requereixi massa temps per a la seva utilització com a eina en estudis d'ecologia microbiana amb moltes mostres. Per altra banda, alguns autors han assenyalat l'obtenció de seqüències de no AOB en utilitzar amoA en un protocol de PCR-DGGE.
Amb la finalitat d'obtenir una eina ràpida i rigorosa per detectar i identificar els AOB, vam desenvolupar un joc nou d'oligonucleòtids amb diana en el gen amoB, que codifica per a la subunitat transmembrana de l'enzim AMO. Aquest gen ha demostrat ser un bon marcador molecular pels AOB, oferint, sense tenir en compte afiliacions filogenètiques, una elevada especificitat, sensibilitat i fiabilitat.
En aquest treball també presentem una anàlisi de RT-PCR basada en la detecció del gen amoB per a la quantificació del gènere Nitrosococcus. El nou joc d'oligonucleòtids dissenyat permet una enumeració altament específica i sensible de tots els γ-Nitrosococcus coneguts.
Finalment, vam realitzar un estudi poligènic, comparant i avaluant els marcadors amoA, amoB i 16SrDNA, i vàrem construir un arbre filogenètic combinat.
Com a resultat concloem que amoB és un marcador adequat per a la detecció i identificació dels AOB en mostres ambientals, proporcionant alhora agrupacions consistents en fer inferències filogenètiques. Per altra banda, la seqüència sencera del gen 16S rDNA és indicada com a marcador en estudis amb finalitats taxonòmiques i filogenètiques en treballar amb cultius purs de AOB.
Human activities such as farming and industrialization have produced a significant increase in the number of ammonium-rich environments. The presence of nitrogenated compounds reduces water quality causing toxicity problems, deteriorating the environment and even affecting human health. Consequently, nitrification has recently become a widespread process involving the cycling of nitrogen in the biosphere, which is mainly due to microbial activities. Ammonia oxidizing bacteria (AOB) are an essential component of the global cycling of nitrogen, being responsible for the aerobic oxidation of ammonium to nitrite.
Although the first ammonia oxidizers were isolated by the end of the XIX century, the slowness of their growth and the difficulties in culturing hindered achieving a full knowledge of this bacterial group until the 80s, when the first studies based on the gene 16S rDNA where performed. Nowadays, the databases contain huge numbers of entries of 16SrDNA sequences belonging to AOB.
The aim of this work was to find, develop, and evaluate useful and reliable tools for the study of ammonia oxidizers in environmental samples.
In this work we describe the use of Fluorescence In Situ Hybridization (FISH), based on the use of DNA probes specifically targeting the ammonia-oxidizers 16SrRNA molecule. AOB were detected and enumerated by using this technique. However, unknown sequences are hardly detectable by using this method, and therefore, new tools were needed.
For this purpose we tried applying the sequence of the probe Nso1225 in a PCR reaction. The possibility of specifically amplifying a 16S rDNA gene fragment resulted in a new fingerprinting tool to assess the presence and diversity of ammonia-oxidizers in natural environments. Even so, some β-Proteobacterial non-AOB sequences were also retrieved by using this technique. Moreover, one of the main disadvantages of using 16S rDNA as a molecular marker is the impossibility of simultaneously detecting both the β and the γ-Proteobacterial ammonia oxidizers.
The gene amoA, which encodes for the subunit A of the enzyme ammonia monooxygenase, was then being extensively used as a marker for the detection of AOB in environmental samples. We describe the use of this marker for the identification of several ammonia oxidizing sequences in sludge samples from a sequencing batch reactor (SBR). Although useful, the use of amoA as a marker requires cloning, which is a tedious and time-consuming technique when dealing with large number of samples in microbial ecology studies. Besides, detection of non-AOB sequences has been reported by other authors when using amoA in a PCR-DGGE approach.
Aiming at obtaining a fast and rigorous analytical tool allowing AOB detection and identification, we developed a new set of primers targeting the gene amoB, which encodes for the transmembrane domain of the enzyme ammonia monooxygenase. This gene has been shown to be a good molecular marker for AOB, since it can be used for easy detection and identification of ammonia oxidizers, providing high specificity, sensitivity and reliability regardless of phylogenetic affiliations.
A real-time PCR assay for the detection and quantification of the γ-proteobacterial genus Nitrosococcus based on the amoB gene sequence is also presented. This newly designed primer set allows a highly sensitive and specific enumeration of all known Nitrosococci.
We finally performed a comparison and evaluation of the markers amoA, amoB and 16S rDNA, and built a polygenic based tree.
As a result we conclude that amoB is a suitable molecular tool for detecting and identifying AOB in environmental samples, yielding consistent grouping when performing phylogenetic inferences. In turn, the whole sequence of the gene 16S rDNA is indicated for taxonomical and phylogenetic purposes when working with ammonia oxidizing isolates.
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Petry, Frauke. "Charakterisierung eines neuen ATP-binding-cassette-Transporters aus der ABCA-Subfamilie." Doctoral thesis, [S.l.] : [s.n.], 2004. http://webdoc.sub.gwdg.de/diss/2004/petry/petry.pdf.
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Xu, Meng. "Specialised transcription factories." Thesis, University of Oxford, 2008. http://ora.ox.ac.uk/objects/uuid:a41d3243-c233-491a-916b-4e329cace434.
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The intimate relationship between the higher-order chromatin organisation and the regulation of gene expression is increasingly attracting attention in the scientific community. Thanks to high-resolution microscopy, genome-wide molecular biology tools (3C, ChIP-on-chip), and bioinformatics, detailed structures of chromatin loops, territories, and nuclear domains are gradually emerging. However, to fully reveal a comprehensive map of nuclear organisation, some fundamental questions remain to be answered in order to fit all the pieces of the jigsaw together. The underlying mechanisms, precisely organising the interaction of the different parts of chromatin need to be understood. Previous work in our lab hypothesised and verified the “transcription factory” model for the organisation of mammalian genomes. It is widely assumed that active polymerases track along their templates as they make RNA. However, after allowing engaged polymerases to extend their transcripts in tagged precursors (e.g., Br-U or Br-UTP), and immunolabelling the now-tagged nascent RNA, active transcription units are found to be clustered in nuclei, in small and numerous sites we call “transcription factories”. Previous work suggested the transcription machinery acts both as an enzyme as well as a molecular tie that maintains chromatin loops, and the different classes of polymerases are concentrated in their own dedicated factories. This thesis aims to further characterise transcription factories. Different genes are transcribed by different classes of RNA polymerase (i.e., I, II, or III), and the resulting transcripts are processed differently (e.g., some are capped, others spliced). Do factories specialise in transcribing particular subsets of genes? This thesis developed a method using replicating minichromosomes as probes to examine whether transcription occurs in factories, and whether factories specialise in transcribing particular sets of genes. Plasmids encoding the SV40 origin of replication are transfected into COS-7 cells, where they are assembled into minichromosomes. Using RNA fluorescence in situ hybridisation (FISH), sites where minichromosomes are transcribed are visualised as discrete foci, which specialise in transcribing different groups of genes. Polymerases I, II, and III units have their own dedicated factories, and different polymerase II promoters and the presence of an intron determine the nuclear location of transcription. Using chromosome conformation capture (3C), minichromosomes with similar promoters are found in close proximity. They are also found close to similar endogenous promoters and so are likely to share factories with them. In the second part of this thesis, I used RNA FISH to confirm results obtained by tiling microarrays. Addition of tumour necrosis factor alpha (TNF alpha) to human umbilical vein endothelial cells induces an inflammatory response and the transcription of a selected sub-set of genes. My collaborators used tiling arrays to demonstrate a wave of transcription that swept along selected long genes on stimulation. RNA FISH confirmed these results, and that long introns are co-transcriptionally spliced. Results are consistent with one polymerase being engaged on an allele at any time, and with a major checkpoint that regulates polymerase escape from the first few thousand nucleotides into the long gene.
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Coma, Bech Marta. "Biological nutrient removal in SBR technology: from floccular to granular sludge." Doctoral thesis, Universitat de Girona, 2011. http://hdl.handle.net/10803/32025.
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Biological nutrient removal has been studied and applied for decades in order to remove nitrogen and phosphorus from wastewater. However, more anthropogenic uses and the continued demand for water have forced the facilities to operate at their maximum capacity. Therefore, the goal of this thesis is to obtain more compact systems for nutrient removal from domestic wastewater. In this sense, optimization and long-term stabilization of high volume exchange ratios reactors, treating higher volumes of wastewater, have been investigated. With the same target, aerobic granular sludge was proposed as a reliable alternative to reduce space and increase loading rates in treatment plants. However, the low organic loading rate from low-strength influents (less than 1 Kg COD•m-3d-1) results in slower granular formation and a longer time to reach a steady state. Because of that, different methodologies and operational conditions were investigated in order to enhance granulation and nutrient removal from domestic wastewater.L’estudi de l’eliminació biològica de nutrients s’ha dut a terme durant dècades. Tot i això, la influencia de l’home i l’augment de la demanda d’aigua han forçat a les instal•lacions a treballar a la seva capacitat màxima. Així, l’objectiu de la tesi és obtenir sistemes més compactes per a l’eliminació de nutrients de les aigües residuals. En aquest sentit, s’ha investigat l’optimització i estabilització de reactors amb alts volums d’intercanvi, tractant més aigua. Amb el mateix objectiu, el fang granular aeròbic va ser proposat com una alternativa fiable per tal de reduir l’espai i incrementar les càrregues de les depuradores. Tot i això, la granulació amb influents de baixa càrrega (menors a 1 Kg dQO•m-3d-1) resulta més lenta i més dificultosa alhora d’obtenir l’estat estacionari. Per aquesta raó es van investigar diferents metodologies i condicions d’operació per tal de millorar la granularció i l’eliminació de nutrients de les aigües urbanes.
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Anamani, Denise E. "A fluorescence in situ hybridization (FISH) analysis of human lung cancer /." 2004. http://www.consuls.org/record=b27074626.
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Thesis (M.A.)--Central Connecticut State University, 2004.Thesis advisor: Kathy Martin-Troy. " ... in partial fulfillment of the requirements for the degree of Master of Arts in Biology." Includes bibliographical references (leaves 21-23). Also available via the World Wide Web.
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Friedman, Brett. "Characterisation of a chromosomal translocation in an ovarian carcinoma cell line using fluorescence 'in situ' hybridisation." Thesis, 1996. http://hdl.handle.net/10539/22288.
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A dissertation submitted to the Faculty of Health Sciences, University of the Witwatersrand, Johannesburg in fulfilment of the requirements for the degree of Master of Science (Haematology)The region Ilpl3-pl5 on the short arm of chromosome 11 (lip) has been implicated in the initiation or progression of several human malignancies including the embryonic rhabdomyosarcoma Wilms' tumour, bladder, renal cell and ovarian carcinoma. In this study, Fluorescence In Situ hybridisation (FISH) was used to identify the nature of a chromosome llp+ abnormality present in two ovarian carcinoma cell lines after conventional cytogenetic techniques had failed to elucidate the chromosomal origin of the abnormality. Using whole chromosome library probes, the abnormality in cell line UW0V2 was found to be composed entirely of chromosome 3 material representing the translocation t (3;11) (pl2-14;pl5). In the protein-free subline UW0V2(Sf), the abnormality was found to consist of the complex translocation t (3;8; 11) (pl2-14 ;q22-24;pl5) . It is possible that the involvement of chromosome 8 in this translocation was a cell culture phenomenon. Other structural and numerical abnormalities elucidated with FISH in cell line UW0V2(Sf) included lq+, +5, +7, 7q-, 8q+, +12, +14, 14q+, -15, 16q- and -18. Using FISH together with the gene probe pSB|5 and the CEPH YAC probes 892g9, 785e5, 847al2, 954f4, 966e8 and 845a3, the breakpoint region on chromosome 11 in the _ two cell lines was narrowed down and mapped to the region Ilpl4.3-pl5.1 lying between probes 966e8 (D11S902) and 845a3 (D11S899). This represents a physical distance of approximately 1 Mb. The breakpoint in the two cell lines appeared to involve the same region on llplS.l. In a separate study, three epithelial ovarian tumour specimens and four ascitic fluid specimens were obtained. Tumour specimens T2 and T4 and ascitic fluid specimens AF-1, AF-2 and AF-3 were all cytogenetically uninformative. Cytogenetic analysis of specimen T5 revealed a single clonal abnormality involving a deletion in the region 6q21. Ascitic fluid specimen AF-5 yielded cytogenetically normal metaphases. Both specimens were hypodiploid and revealed a cytogenetically normal chromosome 11. Using FISH and CEPH YAC probes 966e8 and 845a3, no abnormalities were detected in the region llpl4.3-pl5.1 in these two specimens but one cannot rule out the possibility of submicroscopic abnormalities lying within the region between these probes. From this study we speculate that chromosome 6 abnormalities may be important in the initiation of these tumours. From the results obtained with cell lines UW0V2 and UW0V2 (Sf) we speculate that the chromosome 3 abnormalities were an early event in the evolution of these tumours while the chromosome 11 abnormality was a later event. Little is known about the region llpl4.3-pl5.1 and very few disease loci have been assigned to this region, however, we may speculate that this region harbours a tumour suppressor gene or an oncogene whose disruption or activation is critical to the pathophysiology of ovarian carcinoma and other genitourinary cancers.
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Swaneburg, Uwe [Verfasser]. "Detektion von Porphyromonas gingivalis mittels Fluoreszenz-in-Situ-Hybridisation (FISH-Technik) / vorgelegt von Swaneburg, Uwe, geb. Patzer." 2006. http://d-nb.info/992236258/34.
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