magistrsko delo
Maja Zugan (Author), Darja Keše (Reviewer), Darja Žgur-Bertok (Mentor)

Abstract

Uropatogene Escherichia coli (UPEC) povzročajo okužbe sečil. Ključno vlogo pri tem imajo virulentni dejavniki, med katerimi najpogosteje zasledimo fimbrije, toksin hemolizin ter sisteme za privzem železa. Izražanje genov za virulentne dejavnike bi lahko bilo podvrženo heterogenosti, pojavu ki ga zasledimo v populaciji genetsko enakih celic in je posledica okoljskih dejavnikov, ki vplivajo na razlike v translaciji, transkripciji in topologiji DNA. Da bi natančneje preučili izražanje genov za virulentne dejavnike na ravni posameznih celic, smo na plazmidnem vektorju pBAC, spojili promotorje genov za fimbrije tipa 1 (fimA), ? hemolizina (hlyA) in za kelator železa, aerobaktin (iucD) z genom za fluorescentni protein Gfp brez lastnega promotorja. Na izražanje preiskovanih promotorjev smo s fluorescentno mikroskopijo spremljali vpliv hranil, pomanjkanja železa, pH in rast na trdni podlagi. Ugotovili smo, da sta se promotorja genov iucD in hlyA izražala v višji meri na minimalnem gojišču s kelatorjem železa dipiridilom, medtem ko smo za promotor gena za fimbrije fimA pokazali višjo raven izražanja v hranilnem gojišču LB. Izražanje vseh treh preiskovanih promotorjev je bilo načeloma homogeno, vendar smo zasledili primer heterogenega izražanja promotorja iucD v pogojih, ko izražanje sicer ni bilo inducirano.

Keywords

Escherichia coli;hemolizin;fimbrije;aerobaktin;izražanje genov;heterogenost;fluorescenca;

Data

Language: Slovenian
Year of publishing:
Typology: 2.09 - Master's Thesis
Organization: UL BF - Biotechnical Faculty
Publisher: [M. Zugan]
UDC: 579.25:579.842.1/.2:577.2
COBISS: 5088888 Link will open in a new window
Views: 596
Downloads: 137
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Other data

Secondary language: English
Secondary title: Expression of selected virulence factors genes in bacteria Escherichia coli
Secondary abstract: Uropathogenic Escherichia coli (UPEC) are a leading cause of urinary tract infections and virulence factors play a key role in pahogenesis - fimbriea, toxins such as hemolysin and iron uptake systems such as aerobactin, being the most common. The expression of virulence factor genes might be heterogenic, meaning that all genetically identical cells in a population, do not express the same genes. It is belived that heterogeneity is affected by differences in rates of translation, transcription and DNA topology. To further understand the expression of virulence factor genes we constructed gene fusions using the pBAC plasmid, the gfp gene for the Gfp fluorescent protein without its promoter and fimA, hlyA and iucD promotors for fimbriae, hemolysin and aerobactin, respectively. Using fluorescent microscopy and a variety of different growth conditions we were able to study gene expression at a single cell level. The employed growth media were: rich LB medium, minimal media with an added iron chelating ligand, as well as a low pH medium, all in liquid and firm form. Results showed higher promotor iucD expression when cells were grown on minimal media with iron deficiency, whereas the fimA fimbriae promotor showed higher expression upon growth on rich medium. In principle, expression was found to be homogenous however, an example of heterogenous expression of iucD was observed under noninducing conditions.
Secondary keywords: Escherichia coli;hemolysin;fimbriae;aerobactin;gene expression;heterogeneity;fluorescence;
Type (COBISS): Master's thesis/paper
Study programme: 0
Thesis comment: Univ. v Ljubljani, Biotehniška fak., Študij mikrobiologije
Pages: X, 45 f.
ID: 11215336