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Optimalizace postupů pro kvantifikaci miRNA z tenkojehlových bioptických vzorků karcinomu pankreatu. / Optimization of miRNA analysis in fine-needle biopsy samples of pancreatic cancer tissue.Čuperková, Romana January 2014 (has links)
Pancreatic cancer (PC) is extremely severe malignant disease with a five-year survival of less than 5%. Currently there is no reliable tool for the diagnosis of PC in its early stages. At the time of clinical symptoms most patients are in an advanced stage of the disease and the treatment does not usually have a significant effect. For these reasons emphasis is gradually shifting to the search for the suitable molecular markers for improvement of the diagnosis and assessment of the survival prognosis with respect to a possibility of surgical treatment. MiRNA represent one of the most promising markers, although, their examination in pancreatic tissue is a complicated process. One of the reasons is the very small amount of the source material coming from a fine needle biopsy. A second cause of problems is the subtle character of the pancreatic tissue resulting in significantly lower yields of molecular genetic analysis when compared to other epithelial tissues. An additional negative factor is heterogeneity of the tissue resulting in disproportionate representation of tumor cells within the sample. A suitable choice of procedures for isolation of nucleic acids (NA) and subsequent analysis including quantification of tumor cells is critical for accurate evaluation of the miRNA levels. This work is...
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Detection And Characterization Of Plant Genes Involved In Various Biotic And Abiotic Stress Conditions Using Ddrt-pcr And Isolation Of Interacting ProteinsUnver, Turgay 01 August 2008 (has links) (PDF)
The main objective of this thesis dissertation is functionally characterizing the genes involved in biotic and abiotic stresses of plants at molecular level. Previously, upon pathogen attack Rad6 gene expression was found to be changed in wheat and barley plants. To functionally characterize the Rad6 gene, VIGS (Virus induced gene silencing) system was used. HR (Hypersensitive response) like symptoms was detected in every silenced barley and wheat plants. To figure out, transcriptomes and proteomes of Rad6 silenced plants were analyzed. 2-D PAGE analysis was also performed on silenced and control wheat plants. No pathogen growth was observed in Rad6 silenced barley lines. Additionally, the susceptible wild type Arabidopsis plants showed resistant phenotype when any of the Rad6 gene copies is mutated. This suggests that Rad6 gene has a negative regulatory role in plant disease resistance which was proved for the first time. Yeast two hybrid protein interaction study suggests that RAD6 carrying out its function by interacting with SGT1 protein and regulating resistance related genes. It has been first time reported in this thesis that E2 (Ubiquitin conjugating enzyme) takes role in plant disease resistance.
Boron which is the other consideration in the scope of thesis as an abiotic stress factor at a very limited amount is necessary for the normal development of plants. This study is conducted on highly boron tolerant Gypsophila perfoliata L. collected from a location in the boron mining area. The plant samples were tested in the presence of high boron (35 mg/kg) concentrations. The transcriptomes of the plant samples treated with the excess levels of boron to that of the samples grown under normal concentration were compared using differential display PCR method. Thirty bands showing differential expression levels at varying time points were analyzed. 18 of them were confirmed via qRT-PCR.
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Elucidation Of The Role Of Gcn2 Gene In Response To Powdery Mildew InfectionOzturk, Ibrahim Kutay 01 August 2012 (has links) (PDF)
Plant immune system is entirely based on the immunities of the individual cells in which systemic signals originate from the infection sites. Powdery mildew disease is one of the agents causing these infection sites, resulting in significant yield losses, if disease develops. Understanding the molecular basis of plant-pathogen interactions is the new trend for fighting against plant pathogens, since classical methods used in selection of resistant plants are becoming less and less efficient nowadays. Thus, finding out the genes which are responsible in plant&rsquo / s resistance is becoming very important.
In this thesis, effect of &lsquo / General Control Nondepressible-2&rsquo / (GCN2) homolog protein in barley defense mechanism was aimed to be studied. The GCN2 of yeast was
v
previously identified in our laboratory as an interacting protein when the yeast cDNA library was screened with a putative yellow rust R gene (Yr10) fragment. There are reports available in the literature for the function of GCN2 protein, which makes it a good candidate for a role in disease resistance. Thus, the barley homologue of GCN2 might have a role in the R protein mediated early disease response of which may be proceeding via Programmed Cell Death (PCD). In order to observe such function of HvGCN2 in barley, silencing of its expression via Virus Induced Gene Silencing (VIGS) was investigated. Therefore, the GCN2 homologue was found to function as dampening the severity of the disease.
The silencing with triple technical replicates was observed in 5 of the 6 samples, at an average of 43.2% by qRT-PCR analysis. The pathogen growth levels at different time points were analyzed under light microscope on the silenced and the control samples by measuring the primary and secondary hyphae lengths. The total of 24 seedlings and 292 individual spores were analyzed, and then the level of disease formation was quantitated with 603 primary hyphae and 106 secondary hyphae measurements. Up to 25% hyphae growth rate differences between the control and silenced groups were observed with a probability value less than 0.05 on t-test.
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Stress and the Offspring : Adaptive Transgenerational Effects of Unpredictability on Behaviour and Gene Expression in Chickens (<em>Gallus gallus</em>)Nätt, Daniel January 2008 (has links)
<p>Environmental stress has shown to affect both the exposed individuals and the development of their offspring. Generally, it is thought that the stressed organism responds to stress by trying to adapt to it. This thesis investigates possible evolutionary consequences of cross-generational transmissions of stress, where the parent has been stressed but the offspring has not. In two studies we have exposed chicken parents of different breeds to an unpredictable circadian light rhythm, to investigate the influence of genetic background on the transmission of behaviour and patterns of genome-wide gene expression across generations. In Paper I, we can show that the domesticated chicken, by means of epigenetic factors, transmit their behaviours as well as their gene expression profiles to their offspring to a higher extent than their wild ancestor, the red junglefowl. Furthermore, in Paper II, even though the offspring never experienced the stress or had any contact with their stressed parents, they seemed to have adapted to it, which suggests that the parents might have prepared (or pre-adapted) them for living in the unpredictable environment. Additionally, eggs of stressed hens showed increased levels of estradiol that might have affected gene expression of specific immune genes, which were up-regulated in the offspring of stressed parents. It is possible that the traditional distinction between stress responses and evolutionary adaptation may be reevaluated, since our results indicate that they could be parts of the same evolutionary event.</p>
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The molecular characterization of South African isolates of Grapevine Rupestris Stem Pitting-associated virus (GRSPaV)Noach, Liesl Christine 12 1900 (has links)
Thesis (MSc (Genetics))--University of Stellenbosch, 2010. / Includes bibliography. / ENGLISH ABSTRACT: The first aim of this study was to reliably and rapidly detect Grapevine rupestris stem pittingassociated
virus (GRSPaV) in grapevine. This was achieved by screening 94 grapevines
using crude plant extracts in both quantitative and conventional reverse transcription
polymerase chain reaction (RT-PCR). The second aim was to establish a technique capable of
differentiating GRSPaV sequence variants. The application of this technique is for the largescale
screening of diseased vines to associate sequence variants of GRSPaV with disease
symptoms. Nested quantitative polymerase chain reaction and high resolution melting assays
(qPCR-HRM) were developed for three regions of the GRSPaV genome (coat protein, RNAdependant
RNA-polymerase and triple gene block movement protein). The qPCR-HRM
technique using the high saturation dye, EvaGreen™, and the Rotor-Gene™ 6000 analyzer
was validated with a panel of sixteen sequence-characterized viral isolates. Diluted RT-PCR
products and cloned cDNA gave the most consistent amplification plots and dissociation
profiles. RT-PCR products generated from total RNA extracts were used as template for
qPCR-HRM assays and for direct sequencing of sixteen samples in the three aforementioned
regions. The average amplification efficiency for qPCR was 1.52±0.04. Auto-calling of userdefine
genotypes was performed at a confidence interval of 70%. Phylogenetic analysis of the
three regions of the GRSPaV genome was performed with published GenBank sequences to
confirm the HRM data. The dominant sequence variants found in the South African sample
set radiated with Group II, reference full-length variant GRSPaV-SG1. GRSPaV-infected
samples can in future be subjected to qPCR-HRM assays developed during this study. This
can be performed to establish similarity to known genotypes and therefore phylogenetic
groups. Mixed infection of sequence variants and quasi-species were a common occurrence.
The assay will be useful in establishing correlation of specific genotypes to different
phenotypical expression of viral disease. This could provide insight into the etiology of
diseases associated with GRSPaV. / AFRIKAANSE OPSOMMING: Die eerste doel van hierdie studie was om die virus wat met Rupestris-stamverpitting
(Grapevine rupestris stem pitting-associated virus of “GRSPaV”) in wingerd verbind is,
vinnig en betroubaar op te spoor. Dit is bereik deur 94 wingerdstokke vir die
teenwoordigheid van die virus te toets met beide kwantitatiewe en konvensionele trutranskripsie
polimerase kettingreaksies (RT - PCR) vanaf ongesuiwerde plant-ekstraksies.
Die tweede doel was die daarstelling van ’n tegniek om onderskeid te tref tussen variante van
GRSPaV met verskillende nukleotiedvolgordes. Hierdie tegniek kan op groot skaal gebruik
word om ge-affekteerde wingerdstokke te toets om sodoende siektesimptome met spesifieke
variante van GRSPaV te verbind. Ge-neste kwantitatiewe polimerase-kettingreaksies (qPCR)
en hoë-resolusie smelt-analises (HRM) is ontwikkel vir drie streke van die GRSPaV-genoom
(mantelproteïen, RNS-afhanklike RNS-polimerase en trippelgeenblok bewegingsproteïen).
Die tegniek van qPCR-HRM met die hoë-versadingingskleurstof EvaGreen™ en die Rotor-
Gene™ 6000 ontleder se geldigheid is bevestig deur vergelyking met ’n paneel van sestien
virus-isolate waarvan die volgorde reeds bepaal is. Verdunde RT-PCR-produkte en
gekloneerde DNS het die mees konsekwente amplifikasie-uitstipping en dissosiasieprofiele
opgelewer. RT-PCR-produkte wat vanuit totale RNS-ekstrakte verkry is, is as templaat vir
qPCR-HRM-analises gebruik. Dieselfde produkte is ook gebruik, om die volgorde van
sestien monsters in drie streke direk te bepaal. Die gemiddelde amplifikasiedoeltreffendheid
van die qPCR was 1.52±0.04. Gebruiker-gedefinieerde genotipes is deur middel van outooproeping
teen ’n vertroue-interval van 70% uitgevoer. Filogenetiese analises vir drie streke
van die GRSPaV-genoom is uitgevoer met gepubliseerde GenBank-volgordes om die HRMdata
te bevestig. Die dominante volgorde-variante in die stel Suid-Afrikaanse monsters het
ooreengestem met Groep II, vollengte-verwysingsvariant GRSPaV-SG1. Monsters wat met
GRSPaV besmet is kan in die toekoms onderwerp word aan die qPCR-HRM-analises wat in
hierdie studie ontwikkel is. Dit kan uitgevoer word om ooreenkomste met bekende genotipes
te bepaal, en dus ook met filogenetiese groepe. Die besmetting van plante met meer as een
volgorde-variant het algemeen voorgekom. Die kwasi-spesies populasie-struktuur van die
virus het ook gedurig na vore gekom. Die toets sal nuttig wees in die bepaling van korrelasies
tussen spesifieke genotipes en verskillende fenotipiese voorkomste van virussiektes. Dit kan
insig verleen in die etiologie van siektes wat met GRSPaV verbind word.
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Analyse transcriptomique de deux souches fongiques québécoises Inonotus obliquus et Armillaria sinapinaFradj, Narimane January 2019 (has links) (PDF)
No description available.
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Topical Black Raspberries and Strawberries Bioincorporated with Selenium Reduce Experimental Oral CancerWarner, Blake Matthew 26 August 2013 (has links)
No description available.
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Morfologická a funkční charakterizace střevního epitelu z hlediska exprese proteinu LGR4 / Morphological and functional characterization of intestinal epithelium in the context of LGR4 expressionBurešová, Petra January 2017 (has links)
No description available.
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