Mastergradsoppgaver i molekylær bioteknologi
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Mastergradsoppgaver i molekylær bioteknologi
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Nguyen, Lam Van (Master thesis; Mastergradsoppgave, 24-Dec-2008)[+][-]
Abstract: Alpha-mannosidosis is a rare autosomal recessive lysosomal-storage disorder resulting from deficiency of lysosomal alpha-mannosidase activity. The disease shows a wide range of clinical phenotypes caused by intracellular accumulation of mannose-containing oligosaccharides, which ultimately may lead to mental retardation, hearing loss, skeletal changes and immune deficiency. Lysosomal alpha-mannosidase is a hydrolase that cleaves alpha-linked mannose residues from the non-reducing end of N-linked glycans of glycoproteins. So far, 40 alpha-mannosidosis associatied sequences have been reported, but no correlation of clinical and disease associated sequence variations has been detected so far. In previous work, the seven alpha-mannosidosis associated mutations, p.C55F,p.H200L, p.P263L, p.S318L, p.S453F, p.S457E and p.T745R in the LAMAN gene were identified. In order to characterize the functional of these sequence variants at the biochemical and cellular levels to investigate if they are disease-causing mutations. The mutations were introduced into an expression vector containing the wild-type LAMAN cDNA by in vitro mutagenesis, and the resulting proteins were expressed in COS-7 and BHK cells. This study had revealed that the previous indentified mutation p.C55F suggested to cause human alpha-mannosidosis result in no enzyme significant activity in transfected COS-7 and BHK cells. This missense-mutation probably resulted in misfolding and ER-retention. This was consistent with the low enzyme expression level and non lysosomal localization in transfected cells. The six other mutant variants resulted either in no significant residual or low residual activity, but they were intracellularly proteolytically processed and partially sorted to the lysosomes similar to wild-type. URI: http://hdl.handle.net/10037/2413 File(s) in this item: 1
thesis.pdf (2.586Mb) -
Lea, Ingrid (Master thesis; Mastergradsoppgave, 01-Jun-2008)[+][-]
Abstract: Helicobacter pylori is a Gram-negative flagellated bacterium that infects the stomach of around 50 % of the worlds population, and can give rise to serious disease such as chronic gastritis, duodenal ulcer and stomach cancer. The current treatment against H. pylori, consisting of a combination of a proton pump inhibitor and two different antibiotics, is effective but has several drawbacks such as increased risk of antibiotic resistance and no protection against re-infection. Although extensive research is going on to develop a vaccine that would be an attractive alternative or complement to the current treatment, there is no vaccine available against H. pylori today. Antibodies to a number of antigens have been detected in the sera of infected patients, and many of these are known to be virulence factors, and are considered as candidate H. pylori vaccine antigens, alone or in a combination. Two such antigens are Neutrophil-activating protein (NAP) and Flagellin A (FlaA). In this project the H. pylori antigens NAP and FlaA were cloned and expressed in E. coli, and a purification strategy were set up for the two proteins. URI: http://hdl.handle.net/10037/1650 File(s) in this item: 2
thesis.pdf (5.971Mb)vedlegg.pdf (387.1Kb) -
Phung, Danh Thanh (Master thesis; Mastergradsoppgave, 01-Jun-2008)[+][-]
Abstract: To understand the regulation of the pathophysiological processes, such as inflammation, thrombosis and atherosclerosis, it is very important to characterize the interactions between circulating cells and which molecules that contributes to and promote these interactions. As a part of this, the role of eicosanoids in cell-cell interactions in whole blood ex vivo and isolated blood cells are investigated. In this study, various inhibitors were used to regulate the amount of prostaglandins (and leukotrienes) and how these eicosanoids affect the activity- and expression level of tissue factor (TF), cytokines, enzymes, and receptors involved in the intercellular communication. Before the inhibition study, time course experiments revealed that the incubation times between 1.5 h and 2 h was sufficient for further study of the parameters under investigation. In the inhibition study, whole blood samples were preincubated with different eicosanoid inhibitors, and then stimulated with LPS for ninety minutes prior to TF activity measurement and real-time PCR analysis of cytokine gene expressions. Aspirin did not significantly enhance the lipopolysaccharide (LPS)-induced TF activity in whole blood, however a trend for enhance induction was indicated. URI: http://hdl.handle.net/10037/1648 File(s) in this item: 1
thesis.pdf (1.457Mb) -
Nyrud, May Liss Julianne (Master thesis; Mastergradsoppgave, 01-Jun-2008)[+][-]
Abstract: The marine fish pathogen Vibrio salmonicida is the causative agent for vibriosis in Atlantic salmon (Salmo salar L.), Rainbow trout (Oncorhynchus mykiss) and Atlantic cod (Gadus morhua L.). V. salmonicidas virulence is regulated by Quorum sensing (QS) systems, which includes important regulatory small RNAs (sRNAs). sRNAs have the last years been identified in large numbers, and mostly in pathogenic bacteria strains. A cDNA library from small RNA species (120-340 nt) was constructed by size fractionation on a polyacrylamide gel. Due to time limitations, it was not possible to quality check the library properly. More colonies need to be screened in order to decide whether the cDNA library can be used in further studies or not. One sRNA proven to be very important in QS in several Vibrios is the quorum regulating RNA (Qrr). The expression of qrr has earlier been confirmed by Northern blotting. Furthermore, V. salmonicida has got only one predicted Qrr encoding gene, while other Vibrios have several. In order to gain more information about the role of Qrr in V. salmonicida, the construction of a qrr knock-out by deletion mutant was initiated. The knock-out construct was created, but the conjugation was not successful. The conjugation method for V. salmonicida is not yet established, so further optimization needs to be developed. A cDNA library of high quality can be subject to further experiments including ultra-high throughtput DNA sequencing followed by expression analysis like Northern blot. A qrr knock-out of V. salmonicida can be further investigated by methods like Microarray. Discovery of novel sRNAs can give us new insights in V. salmonicidas growth, virulence and stress adaptation and reveal possible targets for new antibiotics. URI: http://hdl.handle.net/10037/1646 File(s) in this item: 1
thesis.pdf (3.150Mb) -
Henriksen, Hanne F. (Master thesis; Mastergradsoppgave, 08-Jun-2006)[+][-]
Abstract: Many plants in northern regions require a period of low temperatures and short days (called vernalisation) during the winter season to initiate the flowering process. In this experiment, some genes thought to be involved in vernalisation response and induction of flowering in meadow fescue (Festuca pratensis) has been investigated. The genes were chosen based on their possible contribution in the transition from vegetative to reproductive phase in cereals. The investigations were predominantly gene expression studies on vernalised and non-vernalised plant material from genotypes that are able to flower without vernalisation and genotypes with vernalisation requirement. These experiments were done using RT-PCR with cDNA from the different plant genotypes as templates. VRN1, a positive regulator of flowering in cereals, was up regulated by vernalisation in F. pratensis. The results also showed a connection between VRN1 expression and vernalisation requirement. Genotypic differences have been identified in the VRN1 promoter or intron 1 in cereals with differences in vernalisation requirement. These areas are suggested to contain binding sites for repressors of VRN1. The first 1kb of F. pratensis VRN1 intron 1 was sequenced and shown to be identical in the two parents of our mapping population (with different vernalisation requirement). The expression of the putative flowering repressor, MADS16, seemed to be down regulated by vernalisation in the plants that required vernalisation to flower. The plants that were able to flower without vernalisation had a MADS16 expression un-affected by vernalisation. Some investigations were also done on VRN2 (a putative repressor of VRN1 expression), PHYC (a photoreceptor), RUBQ2 (ubiquitin) and CONSTANS (a positive regulator of flowering in Arabidopsis). URI: http://hdl.handle.net/10037/230 File(s) in this item: 2
appendix.pdf (177.2Kb)thesis.pdf (646.0Kb)
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