Priprava in analiza izražanja fuzij genov za virulentne dejavnike bakterije Escherichia coli in genov za fluorescentne proteine

Petrič, Andreja (author), Žgur Bertok, Darja (mentor)

Sevi bakterije Escherichia coli so zelo raznoliki. Okužbe povzročajo sevi, ki so pridobili zapise za sintezo virulentnih dejavnikov. Pri komenzalih in patogenih sevih E. coli so za prvi stik z gostiteljem pomembni adhezini, za preživetje različni sistemi za prevzem železa in izogibanje imunskemu sitemu gostitelja. Patogeni bakterijski sevi imajo tudi zapisi za toksine, ki delujejo na gostiteljeve celice. V populaciji genetsko identičnih bakterijskih celic je poznan fenomen fenotipske variabilnosti. Heterogeno izražanje genov bakterijam nudi prednost v spremenljivem okolju, saj je tako del populacije pripravljen na hitre in neugodne spremembe. Namen naše raziskave je bil pripraviti genske fuzije promotorjev nekaterih genov za virulentne dejavnike in genov za fluorescentne proteine ter analizirati izražanje s fluorescentnim mikroskopom. Preučevali smo promotorje genov, clbA za fosfopanteteinil transferazo, cnf1 za toksin Cnf1 in cdtABC za citotoksin CDT. Genske fuzije smo v več korakih pripravili v plazmidnem vektorju pBAC in ga vnesli v celice bakterijskega seva MG1655. Z genskimi fuzijami smo ugotovili, da vse celice E. coli izražajo gen clbA. Ugotovili smo, da je izražanje genov za sintezo toksina Cnf1 in citotoksina CDT bakterije E. coli heterogeno, saj se izražata le v majhnem delu populacije.

• Language: Slovenian
• Keywords: patogene bakterije, Escherichia coli, virulentni dejavniki, genska fuzija, izražanje genov, fluorescentni proteini, fenotipska heterogenost, fluorescentna mikroskopija
• Work type: Master's thesis/paper
• Typology: 2.09 - Master's Thesis
• Organization: BF - Biotechnical Faculty
• Publisher: [A. Petrič]
• Year: 2017
• PID: 20.500.12556/RUL-96394
• UDC: 579.253.2:579.842:577.21
• COBISS.SI-ID: 4821368
• Publication date in RUL: 30.09.2017

PETRIČ, Andreja, 2017, Priprava in analiza izražanja fuzij genov za virulentne dejavnike bakterije Escherichia coli in genov za fluorescentne proteine [online]. Master’s thesis. A. Petrič. [Accessed 22 June 2025]. Retrieved from: https://repozitorij.uni-lj.si/IzpisGradiva.php?lang=eng&id=96394

Secondary language

• Language: English
• Title: Construction and expression analysis of fusions of Escherichia coli virulence factor genes and genes for fluorescent proteins

Abstract

Escherichia coli is a very diverse bacterial species. Bacterial strains with genes for virulence factors provoke infections. For commensal and pathogenic strains, adhesins are essential for first contact with the host, they also have systems for iron uptake and to avoid the host’s immune system. Pathogenic strains, harbor genes for toxins that damage host cells. In populations of genetically identical bacterial cells, phenotypic heterogeneity has been described. Heterogeneous gene expression provides bacteria an advantage in variable environments and under stress conditions. The purpose of our study was to construct gene fusions of promoters of some virulence factor genes and genes for fluorescent proteins and subsequently analyze their expression with fluorescence microscopy. We studied promoters of genes, clbA for phosphopanteteinyl transferase, cnf1 for toxin Cnf1, and cdtABC for cytoxin CDT. Gene fusions were prepared stepwise in the plasmid vector pBAC and introduced into the bacterial strain MG1655. We found that all cells expressed the clbA gene. Expression of genes for the Cnf1 toxin and cytotoxin CDT was heterogeneous, as they were expressed in only a small subpopulation.

• Keywords: pathogenic bacteria, Escherichia coli, virulence factors, gene fusion, gene expression, fluorescent proteins, phenotypic heterogeneity, fluorescence microscopy