Konjugacija plazmida pOX38:Cm v izbrane seve bakterije Escherichia coli

Repar, Polona-Maja (author), Starčič Erjavec, Marjanca (mentor), Kastrin, Andrej (comentor)

Povečevanje števila proti antibiotikom odpornih bakterij postaja vedno večji problem. Geni za odpornost so pogosto zapisani na mobilnih genetskih elementih, ki se lahko s horizontalnim genskim prenosom hitro prenašajo med bakterijami. Med horizontalnimi genskimi prenosi velja konjugacija, genski prenos ob neposrednem stiku dveh bakterij, kot najpomembnejši dejavnik za širjenje odpornih bakterij. Dobro poznavanje dejavnikov, ki vplivajo na frekvenco konjugacije, lahko vodi v odkritje novih strategij za borbo proti odpornim bakterijam. Med te dejavnike spadajo obrambni mehanizmi bakterij pred vdorom tuje DNA, kot sta na primer sistem CRISPR-Cas in restrikcijsko-modifikacijski (R-M) sistem. Cilj magistrskega dela je bil preveriti, ali obstaja povezava med frekvenco konjugacije plazmida pOX38:Cm in prisotnostjo genskih zapisov za sistem CRISPR-Cas in R-M sisteme v recipientskih sevih, inkompatibilnostjo plazmidov recipientskih in donorskih sevov ali pa s protimikrobnim delovanjem recipientskih sevov. CRISPR-Cas in R-M sisteme smo v posameznih sevih poiskali s prosto dostopnima računalniškima programoma (CRISPR-Cas++ in REBASE), vendar nismo dokazali nobene povezave s frekvenco konjugacije. Nato smo preverili še sposobnost recipienta za zaviranje rasti donorja (test za prisotnost bakteriocinov) in s programom BLAST preverili inkompatibilnost plazmidov v recipientskih sevih s plazmidom pOX38:Cm. Nekateri recipientski sevi so sicer povzročili cono lize donorskega seva HB101 pOX38:Cm, a povezava s frekvenco konjugacije ni obstajala. Primerjava nukleotidnih zaporedij pOX38:Cm in plazmidov recipientskih sevov tudi ni razkrila morebitne inkompatibilnosti med plazmidi. Opaženih razlik v frekvencah konjugacije pOX38:Cm v različne recipientske seve tako nismo mogli povezati ne z sistemom CRISPR-Cas, ne R-M sistemi, ne protimikrobnim delovanjem in tudi ne z inkompatibilnostjo plazmidov.

• Language: Slovenian
• Keywords: plazmidi, frekvenca konjugacije, pOX38:Cm, CRISPR-Cas, restrikcijsko-modifikacijski sistem, inkompatibilnost plazmidov, protimikrobno delovanje
• Work type: Master's thesis/paper
• Typology: 2.09 - Master's Thesis
• Organization: BF - Biotechnical Faculty
• Place of publishing: Ljubljana
• Publisher: [P.-M. Repar]
• Year: 2021
• PID: 20.500.12556/RUL-130413
• UDC: 579.25:577.2.083
• COBISS.SI-ID: 76675843
• Publication date in RUL: 15.09.2021

REPAR, Polona-Maja, 2021, Konjugacija plazmida pOX38:Cm v izbrane seve bakterije Escherichia coli [online]. Master’s thesis. Ljubljana : P.-M. Repar. [Accessed 14 April 2025]. Retrieved from: https://repozitorij.uni-lj.si/IzpisGradiva.php?lang=eng&id=130413

Secondary language

• Language: English
• Title: Conjugation of the pOX38:Cm plasmid into selected bacterial strains of Escherichia coli

Abstract

The growing number of antibiotic resistant bacteria is becoming an increasing problem. The genes for resistance are often carried on mobile genetic elements that can spread rapidly through bacterial populations by horizontal gene transfer. Mechanisms of horizontal gene transfer include conjugation, the gene transfer between two bacteria that are in physical contact, which is considered the most important factor in the spread of resistant bacteria. A good characterisation of the factors that influence the conjugation frequency may lead to the discovery of new strategies to combat the resistant bacteria. These factors include bacterial defence mechanisms to control the entry of foreign DNA, such as CRISPR-Cas and restriction-modification (R-M) systems. The aim of this Master Theses was to determine whether there is a correlation between conjugation frequency of the plasmid pOX38:Cm and the presence of genes for CRISPR-Cas and R-M systems in recipient strains, incompatibility of plasmids in recipient and donor cell or antimicrobial production of recipient strains. CRISPR-Cas and R-M systems were determined using freely available computer programs (CRISPR-Cas++ and REBASE), but a correlation with the conjugation frequency was not discovered. Therefore we investigated the ability of the recipient to inhibit the growth of the donor (bacteriocinogenity assay) and the incompatibility between the plasmids in recipient cells and plasmid pOX38:Cm (BLAST). Some recipient strains formed lysis zones of the donor strain, but a correlation with the conjugation frequency did not exist. Comparison of the plasmids revealed no incompatibility between them. The observed differences in conjugation frequencies could not be explained by CRISPR-Cas or R-M systems, antimicrobial production or plasmid incompatibility.

• Keywords: plasmids, conjugation frequency, pOX38:Cm, CRISPR-Cas, restriction-modification system, plasmid incompatibility, antimicrobial production