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Dissertations / Theses on the topic 'Complementation analysis'
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1
Yilmaz, Seda Aliye. "Complementation Studies To Identify Genes With Roles In Zinc Efficiency In Barley." Master's thesis, METU, 2007. http://etd.lib.metu.edu.tr/upload/12608630/index.pdf.
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Zinc (Zn) is an essential micronutrient for the growth and development of all organisms. Zinc deficiency is a widespread micronutrient disorder worldwide, which reduces crop yields and the nutritive value of the grain. Understanding the process of zinc absorption and translocation in crop is essential for this purpose. Zinc is taken up by plants and translocated within plants through high-affinity zinc transporter proteins embedded in the plasma membrane. The Zn transporters are induced under Zn deficiency, and it is speculated that the expression levels of some of zinc transporters are critical for improved tolerance to low zinc. A number of Zn transporters have been cloned from higher plants including rice and Arabidopsis, but little has been done in barley and wheat. This project aims to investigate genes involveld in zinc efficiency mechanism by complementation analysis in yeast, which is double mutant of zinc-transporters, using cDNA expression library from a most zinc efficient barley cultivar, Tokak-157.
2
Joubert, Jean-François. "A psychomechanical analysis of gerundive and infinitival complementation in intend, mean and propose and their synonyms." Thesis, National Library of Canada = Bibliothèque nationale du Canada, 1997. http://www.collectionscanada.ca/obj/s4/f2/dsk2/ftp04/mq25321.pdf.
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Steinberg, Steven Jeffrey. "Biochemical characterisation and genetic complementation analysis of generalised peroxisomal disorders and Niemann-Pick disease type C." Thesis, King's College London (University of London), 1995. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.294755.
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Mau, Kianna. "Bifluorescent Analysis of ⍺-Synuclein Aggregation In Vivo." Thesis, Université d'Ottawa / University of Ottawa, 2020. http://hdl.handle.net/10393/40937.
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Parkinson’s disease is an incurable neurodegenerative disease characterized by motor deficits, owing to dopaminergic denervation in the nigrostriatal pathway. The abnormal formation of hallmark Lewy bodies underlies the disease process. The pre-synaptic protein alpha- synuclein (⍺-syn) has prion-like properties arising from its propensity to propagate, seed misfolding, and self-aggregate. Pathogenesis is postulated to arise in olfactory and enteric regions, exploiting connected neuronal pathways to ultimately propagate to the substantia nigra pars compacta. There is little known about the earliest stages of ⍺-syn aggregation and its prion-like propagation mechanisms. Bimolecular fluorescence complementation of ⍺-syn aggregates has allowed us to directly visualize aggregation in transgenic mice and mice transduced with an adeno-associated virus vector. Although our transgenic mice expressed BiSyn in a mosaic fashion that limited utility, we were successful in transducing neurons in the mouse striatum. This work has validated the AAV2/9-CMV-BiSyn approach as groundwork for future systematic studies.
5
Wilson, Joyce Anne. "Strategies for the construction and safe release of recombinant baculovirus with enhanced insecticidal properties, intracellular complementation of P74, structural properties of P10 and functional analysis of GP41." Thesis, National Library of Canada = Bibliothèque nationale du Canada, 1997. http://www.collectionscanada.ca/obj/s4/f2/dsk3/ftp04/nq22503.pdf.
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Zhan, Hong. "Analysis of static and dynamic distribution of voltage-dependent calcium channels at nanoscale resolution in Caenorhabditis elegans." Thesis, Paris 6, 2014. http://www.theses.fr/2014PA066333.
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Dans les synapses chimiques, des canaux calciques voltage-dépendants (VDCC) provoquent la fusion des vésicules synaptiques (SV) au niveau de la zone active. L’efficacité et la rapidité de la transmission synaptique dépendent de la distribution relative entre les VDCCs et les SVs prêtes à fusion. Cependant, les modalités d’interaction entre les VDCCs et les SVs ne sont pas connues. Afin de localiser les VDCCs à l’échelle nanométrique j’ai developpé une nouvelle approche chez Caenorhabditis elegans combinant le marquage in vivo des VDCCs, grâce à l’expression d’un épitope extracellulaire, et la microscopie électronique (EM). J’ai généré un transgene GFP::unc-36 qui code la seule sous-unité α2-δ qui s’associe à fois avec les sous-unités formant le pore α1 neuronal (UNC-2) et musculaire (EGL-19) chez C.elegans. J'ai ensuite utilisé des quantum dots conjugués avec l’anticorps anti-GFP, fluorescents et denses au électrons, pour localiser des VDCCs à haute résolution au niveau de la jonction neuromusculaire (NMJ) par EM. En parallèle, j'ai utilisé la technique de CALM (complementation activated light microscopy) pour étudier la dynamique des VDCC dans des vers vivants. Nos résultats montrent que les VDCCs diffusent à l’échelle de nanodomaines sur la membrane musculaire. De plus leur diffusion est modulée en réponse à la tension musculaire. La dystrophine participe au couplage électro-mécanique au niveau du sarcolemme en modulant la taille du domaine de confinement des VDCCs. Enfin, nous avons mis en evidence le rôle de RIM/UNC-10 dans la régulation de la mobilité latérale des VDCCs dans les neurones, probablement via son interaction avec les VDCCs et les SVsAt chemical synapse voltage-dependent calcium channels (VDCC) trigger synaptic vesicles (SV) fusion at the active zone in response to depolarization stimuli. Intracellular Ca2+ influx forms a nanodomain around individual VDCC. Fast and efficient synaptic transmissions appear to be tightly coupled with the relative distribution between the VDCCs and SVs fusion sites. However, the connection between VDCCs and docked SVs at a few nanometer scales remain enigmatic. To localize VDCCs in nanometer resolution I developed a novel approach combining in vivo labeling of VDCCs via genetically-encoded extracellular epitope tags and electron microscopy (EM). I engineered a GFP/split-GFP tag fused at the extracellular N-terminal of UNC-36, the only C. elegans VDCC α2δ subunit associating with both neuronal (UNC-2) and muscular (EGL-19) VDCC pore-forming α1 subunit. I then used quantum dot (QD) conjugated antibodies as both fluorescent and electron dense probes to localize VDCCs at C. elegans neuromuscular junction (NMJ) by in vivo QD-antibodies labeling and EM. In parallel, I applied in vivo complementation activated light microscopy to study VDCC dynamics in live worms. I discovered that VDCCs diffuse within nanodomains at sarcomeric membrane and their nanoscale diffusion behavior is modulated in response to muscle tension. In addition, we found that dystrophin participates in electro-mechanical coupling at the sarcolemma by modulating the confinement size of VDCCs. Meanwhile, we discovered lateral mobility of N-type VDCC at NMJs, and that RIM/UNC-10 seems involved in regulation of VDCC dynamics via its interaction with VDCC and SVs
7
New, Christopher Paul. "Analysis of Tha4 Function and Organization in Chloroplast Twin Arginine Transport." Miami University / OhioLINK, 2020. http://rave.ohiolink.edu/etdc/view?acc_num=miami1586878527570538.
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Jonsson, Rudsander Ulla. "Functional studies of a membrane-anchored cellulase from poplar." Doctoral thesis, KTH, Träbioteknik, 2007. http://urn.kb.se/resolve?urn=urn:nbn:se:kth:diva-4520.
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Cellulose in particular and wood in general are valuable biomaterials for humanity, and cellulose is now also in the spotlight as a starting material for the production of biofuel. Understanding the processes of wood formation and cellulose biosynthesis could therefore be rewarding, and genomics and proteomics approaches have been initiated to learn more about wood biology. For example, the genome of the tree Populus trichocarpa has been completed during 2006. A single-gene approach then has to follow, to elucidate specific patterns and enzymatic details. This thesis depicts how a gene encoding a membrane-anchored cellulase was isolated from Populus tremula x tremuloides Mich, how the corresponding protein was expressed in heterologous hosts, purified and characterized by substrate analysis using different techniques. The in vivo function and modularity of the membrane-anchored cellulase was also addressed using overexpression and complementation analysis in Arabidopsis thaliana. Among 9 genes found in the Populus EST database, encoding enzymes from glycosyl hydrolase family 9, two were expressed in the cambial tissue, and the membrane-anchored cellulase, PttCel9A1, was the most abundant transcript. PttCel9A1 was expressed in Pichia pastoris, and purified by affinity chromatography and ion exchange chromatography. The low yield of recombinant protein from shake flask experiments was improved by scaling up in the fermentor. PttCel9A1 was however highly heterogenous, both mannosylated and phosphorylated, which made the protein unsuitable for crystallization experiments and 3D X-ray structure determination. Instead, a homology model using a well-characterized, homologous bacterial enzyme was built. From the homology model, interesting point mutations in the active site cleft that would highlight the functional differences of the two proteins could be identified. The real-time cleavage patterns of cello-oligosaccharides by mutant bacterial enzymes, the wildtype bacterial enzyme and PttCel9A1 were studied by 1H NMR spectroscopy, and compared with results from HPAEC-PAD analysis. The inverting stereochemistry for the hydrolysis reaction of the membrane-anchored poplar cellulase was also determined by 1H NMR spectroscopy, and it was concluded that transglycosylation in vivo is not a possible scenario. The preferred in vitro polymeric substrates for PttCel9A1 were shown to be long, low-substituted cellulose derivatives, and the endo-1,4--glucanase activity was not extended to branched or mixed linkage substrates to detectable levels. This result indicates an in vivo function in the hydrolysis of “amorphous” regions of cellulose, either during polymerization or crystallization of cellulose. In addition, overexpressing PttCel9A1 in A. thaliana, demonstrated a correlation with decreased crystallinity of cellulose. The significance of the different putative modules of PttCel9A1 was investigated by the construction of hybrid proteins, that were introduced into a knock-out mutant of A. thaliana, and the potential complementation of the phenotype was examined. A type B plant cellulase catalytic domain could not substitute for a type A plant cellulase catalytic domain, although localization and interaction motifs were added to the N- and C-terminus.QC 20100802
9
Weyand, Katrin. "Analysis of the AtMRS2 magnesium transporter family in Arabidopsis thaliana via gene-GFP fusions, heterologous complementations, and protein interaction studies." [S.l.] : [s.n.], 2006. http://deposit.ddb.de/cgi-bin/dokserv?idn=982893264.
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10
Lee, Nagiko Iwata. "Complementation in Japanese : a lexicase analysis." Thesis, 1989. http://hdl.handle.net/10125/9931.
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11
Joubert, Jean-François. "A psychomechanical analysis of gerundive and infinitival complementation in Intend, mean and purpose and their synonyms /." 1997. http://proquest.umi.com/pqdweb?did=738287161&sid=2&Fmt=2&clientId=9268&RQT=309&VName=PQD.
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Day, Shyau-Ming, and 戴學明. "Complementation analysis of the N-domain and the C-domain of the XpsD protein of Xanthomonas campestris pv. campestris." Thesis, 1998. http://ndltd.ncl.edu.tw/handle/98689782170379333662.
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碩士國立中興大學
農業生物科技學研究所
86
XpsD is an outer membrane protein required for the secretion of extracellular enzymes by Xanthomonas campestris pv. campestris. It may serve as the secretion channel by forming multimers. Secondary structure prediction indicated that the protein is rich in β-sheet, except a central region. The XpsD protein may form an N-and a C-terminal β-barrel. Each could serve as an autonomous domain. The purpose of this study is to find out if this hypothesis is correct. Two plasmids were used. One encodes an XpsDN protein and the other encodes an XpsDc protein When the XpsDN. and the XpsDc proteins were co-expressed in the xpsD mutant strain, we observed clear zone surrounding colonies on starch plate. We also detected α-amylase in the extracellular fraction, indicating that the α-amylase was secreted extracellularly. Futhermore, the two proteins cofractionated on gel filtration column. Immunoprecipitation results also suggested that the two proteins may form a stable complex with each other. Overexpression of the XpsDc protein in either wild type or thex xpsD mutant strain inhibited growth. The inhibitory effect was partially relieved by co-expressing the XpsDN protein with the XpsDc protein in the xpsD mutant strain. This result also suggested that the XpsDc protein is no longer inhibitory to the cell growth perthaps as a result of forming functional complex with the XpsDN protein.
13
Hsiao, Po-Yuan, and 蕭柏元. "Analysis of protein-protein interaction of the phosphate homeostatsis-related proteins by using tripartite split-GFP complementation assay in planta." Thesis, 2018. http://ndltd.ncl.edu.tw/handle/7629xd.
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碩士國立清華大學
生物資訊與結構生物研究所
106
ABSTRACT Tripartite split-GFP complementation assay is a new protein-protein interaction technique improved from bimolecular fluorescence complementation (BiFC). Tripartite split GFP system is composed of a larger fragment, GFP1-9 (residues 1–193), and two shorter β-strands : GFP10 (residues 194–212) and GFP11 (residues 213–233). There are some disadvantages of BiFC because BiFC is based on bulky fragments that may increase the difficulty of protein folding and interfere with protein function. To test whether tripartite split-GFP is a useful tool in planta, we first chose two proteins involved in the phosphate starvation reponse: the phosphate starvation response 1 (PHR1) and the SPX domain-containing protein 1 (SPX1) proteins. The PHR1 and SPX1 sandwitch proteins (GFP10-PHR1-GFP11 and GFP10-SPX1-GFP11) were used for transient expression in tobacco leaves and localized in the nucleus. Next, we confirmed the previous results that PHR1 interacts with SPX1 using tripartite split-GFP system. This assay showed that PHR1 and SPX1 can form homo-dimer/oligomers itself. We used the Arabidopsis nitrogen limitation adaptation (NLA/BAH1) to verify the specificity of the interaction between PHR1 and SPX1. Our results showed that NLA interacts with both PHR1 and SPX1. In addition, we chose the other PHR1 family protein PHR1-like 3 (PHL3) to test the interaction of PHL3 with PHR1 and SPX1 respectively. The signal strength between PHR1, SPX1 and PHL3 are different. From the strongest to the weakest is PHR1 and PHL3, PHR1 dimer, SPX1 and PHR1, and then the signal of SPX1 and PHL3 was the strongest. Furthermore, we generated and obtained the Arabidopsis transgenic liens overexpessing GFP10-PHR1, SPX1-GFP11 and GFP1–9. Confonal analysis of these lines showed few signals in the root hair of seedlings, indicating that the tripartite spli-GFP system used in transgenic plants of Arabidopsis needs to be improved.
14
Chen, Yin-Chu, and 陳吟竹. "•Detection of a protein down-regulated in a non-cytotoxic Vibriovulnificus mutant by proteomic analysis •Complementation of a TolC-deficient V. vulnificus strain." Thesis, 2004. http://ndltd.ncl.edu.tw/handle/18425256565599445225.
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碩士國立成功大學
微生物及免疫學研究所
92
Vibrio vulnificus, a gram-negative halophilic marine bacterium, is an opportunistic human pathogen. It causes severe wound infection and septicemia, particularly in those with underlying diseases. V. vulnificus produce a number of potential virulence factors including capsular polysaccharide, siderophores, protease, cytolysin and phospholipase. Our previous study showed that disruption of the genes encoding protease, cytolysin and phospholipase did not influence the virulence in mice or cytotoxicity to HEp-2 cells. Hence we concluded that these three factors are not the major virulence factors in V. vulnificus. However, a non-cytotoxic mutant, NY303, isolated fortuitously from a phospholipase-cytolysin double mutant was shown to be much less virulent in mice. This data suggested that cytotoxicity may be an important virulence factors. We suspected that NY303 might have lost the activity of an unidentified cytotoxin. In this study, we tried to identify the proteins that are lost in NY303 by proteomic analysis. The total soluble proteins, outer membrane proteins, secreted extracellular proteins and periplasmic proteins of NY303 and its isogenic, cytotoxic strain, NY303-2, were extracted and separated by one-dimensional (SDS-PAGE) or two-dimensional electrophoresis (2-DE). When we separated the periplasmic and outer membrane proteins by SDS-PAGE, one band about 35 kDa was found lost in NY303 in each fraction. These bands were subjected to LCQ/MS for protein identities and the transcriptions of genes encoding these proteins were examined by RNA slot blot hybridization. Only with VV2338, the open-reading-frame of one of the identified proteins, the transcript was more abundant in NY303-2 than NY303. However, VV2338 in NY303 can be amplified without any deletion, indicating that the lost of VV2338 protein in NY303 was not caused by a deletion in the gene. Meanwhile, some protein spots differentially expressed in NY303-2 and NY303 were identified on 2-DE gels of the periplasmic and extracellular fraction, suggesting that the lost of cytotoxicity in NY303 might be due to the dysfunction of a regulatory factor. On the other hand, TolC and its homologues are outer membrane proteins essential for the transport of a variety of molecules, including the antibiotics and toxins, across the cell envelope. In our previous study, we found that a TolC-deficient V. vulnificus mutant, MW021, was more sensitive to bile and erythromycin, non-cytotoxic to HEp-2 cells, and much less virulent in mice compared to the parental strain, YJ016. Since the disruption of tolC did not influence the secretion of protease and cytolysin, we suspected that the activity or transport of an unknown cytotoxin was affected by TolC dysfunction. To confirm that the phenotypes we observed were associated with the disruption of tolC, we performed a complementary experiment. We found it’s very difficult to clone the wild-type V. vulnificus tolC gene in Escherichia coli with a high copy number vector, and we always obtained at least one point mutation in the coding sequence of cloned tolC. These suggested that overexpression of V. vulnificus tolC might be harmful for E. coli. When we used the low copy number vector, partial complementation with delayed growth in the 0.02% bile-containing LB medium was observed in one strain, YC019, which contained one point mutation 19-bp upstream of the cloned tolC open reading frame and one silent mutation in the coding sequence. We plated the bacteria grown in the bile-containing LB medium on TCBS (Thiosulfate-Citrate-Bile salt-Sucrose) agar and screened for those that grew as well as YJ016 in bile-containing LB medium. One strain, YC025, was obtained and its phenotypes, including cytotoxicity to HEp-2 cells and susceptibility to erythromycin were similar to YJ016. We found that the tolC gene on the plasmid contained in YC025 had one additional silent mutation and one amino acid deletion. We suspected that the deletion of the amino acid might have altered the TolC structure that resulted in restoration of the phenotypes. Whether TolC has an effect on the activity of an unknown cytotoxin or RTX toxin awaits further investigation.
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Markus, Ralf [Verfasser]. "Complementation of the salt sensitive S. cerevisiae mutant cch1 with an A. thaliana cDNA library and analysis of isolated genes / vorgelegt von Ralf Markus." 2001. http://d-nb.info/963836145/34.
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Chen, Yin-Chu, and 陳吟竹. "‧Detection of a protein down-regulated in a non-cytotoxic Vibrio vulnificus mutant by proteomic analysis ‧Complementation of a TolC-deficient V. vulnificus strain." Thesis, 2004. http://ndltd.ncl.edu.tw/handle/z224j3.
Full textAbstract:
碩士國立成功大學
微生物暨免疫學研究所
92
Vibrio vulnificus, a gram-negative halophilic marine bacterium, is an opportunistic human pathogen. It causes severe wound infection and septicemia, particularly in those with underlying diseases. V. vulnificus produce a number of potential virulence factors including capsular polysaccharide, siderophores, protease, cytolysin and phospholipase. Our previous study showed that disruption of the genes encoding protease, cytolysin and phospholipase did not influence the virulence in mice or cytotoxicity to HEp-2 cells. Hence we concluded that these three factors are not the major virulence factors in V. vulnificus. However, a non-cytotoxic mutant, NY303, isolated fortuitously from a phospholipase-cytolysin double mutant was shown to be much less virulent in mice. This data suggested that cytotoxicity may be an important virulence factors. We suspected that NY303 might have lost the activity of an unidentified cytotoxin. In this study, we tried to identify the proteins that are lost in NY303 by proteomic analysis. The total soluble proteins, outer membrane proteins, secreted extracellular proteins and periplasmic proteins of NY303 and its isogenic, cytotoxic strain, NY303-2, were extracted and separated by one-dimensional (SDS-PAGE) or two-dimensional electrophoresis (2-DE). When we separated the periplasmic and outer membrane proteins by SDS-PAGE, one band about 35 kDa was found lost in NY303 in each fraction. These bands were subjected to LCQ/MS for protein identities and the transcriptions of genes encoding these proteins were examined by RNA slot blot hybridization. Only with VV2338, the open-reading-frame of one of the identified proteins, the transcript was more abundant in NY303-2 than NY303. However, VV2338 in NY303 can be amplified without any deletion, indicating that the lost of VV2338 protein in NY303 was not caused by a deletion in the gene. Meanwhile, some protein spots differentially expressed in NY303-2 and NY303 were identified on 2-DE gels of the periplasmic and extracellular fraction, suggesting that the lost of cytotoxicity in NY303 might be due to the dysfunction of a regulatory factor. On the other hand, TolC and its homologues are outer membrane proteins essential for the transport of a variety of molecules, including the antibiotics and toxins, across the cell envelope. In our previous study, we found that a TolC-deficient V. vulnificus mutant, MW021, was more sensitive to bile and erythromycin, non-cytotoxic to HEp-2 cells, and much less virulent in mice compared to the parental strain, YJ016. Since the disruption of tolC did not influence the secretion of protease and cytolysin, we suspected that the activity or transport of an unknown cytotoxin was affected by TolC dysfunction. To confirm that the phenotypes we observed were associated with the disruption of tolC, we performed a complementary experiment. We found it’s very difficult to clone the wild-type V. vulnificus tolC gene in Escherichia coli with a high copy number vector, and we always obtained at least one point mutation in the coding sequence of cloned tolC. These suggested that overexpression of V. vulnificus tolC might be harmful for E. coli. When we used the low copy number vector, partial complementation with delayed growth in the 0.02% bile-containing LB medium was observed in one strain, YC019, which contained one point mutation 19-bp upstream of the cloned tolC open reading frame and one silent mutation in the coding sequence. We plated the bacteria grown in the bile-containing LB medium on TCBS (Thiosulfate-Citrate-Bile salt-Sucrose) agar and screened for those that grew as well as YJ016 in bile-containing LB medium. One strain, YC025, was obtained and its phenotypes, including cytotoxicity to HEp-2 cells and susceptibility to erythromycin were similar to YJ016. We found that the tolC gene on the plasmid contained in YC025 had one additional silent mutation and one amino acid deletion. We suspected that the deletion of the amino acid might have altered the TolC structure that resulted in restoration of the phenotypes. Whether TolC has an effect on the activity of an unknown cytotoxin or RTX toxin awaits further investigation.
17
Vandas, Karel. "Automatické určování sémantických preferencí pro slovesná valenční doplnění." Master's thesis, 2012. http://www.nusl.cz/ntk/nusl-305141.
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Verb valency plays an important role in the description of behaviour of verbs and connects surface realisation of language with its semantics. Verb itself usually encodes several readings. Complementations of a verb help to identify correct reading of the verb. So far valency verb complementations are mostly studied from morphological and syntactical point of view. The purpose of this thesis is to examine possibilities of automatic identification of semantic preferences for valency complementations of verbs. The thesis discusses performance of system with different levels of available verb valency information in connection with cluster analysis. The thesis contains an evaluation section that compares available methods and their comparision.
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Weyand, Katrin [Verfasser]. "Analysis of the AtMRS2 magnesium transporter family in Arabidopsis thaliana via gene-GFP fusions, heterologous complementations, and protein interaction studies / vorgelegt von Katrin Weyand." 2006. http://d-nb.info/982893264/34.
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Tarasov, Kirill. "Searching for novel gene functions in yeast : identification of thousands of novel molecular interactions by protein-fragment complementation assay followed by automated gene function prediction and high-throughput lipidomics." Thèse, 2014. http://hdl.handle.net/1866/11824.
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