20.500.12556/RUL-180885 Vpliv sorodnosti med bakterijami rodu Bacillus na izmenjavo DNA Influence of relatedness between different Bacillus species on DNA exchange Bacillus subtilis je naravno kompetentna bakterija, ki je sposobna prevzemati zunajcelično DNA iz okolja ter jo integrirati v lasten kromosom. Učinkovitost tega procesa je močno odvisna od stopnje genetske sorodnosti med donorsko DNA in genomom recipienta ter se z njenim zmanjševanjem izrazito zmanjšuje. Poleg tega je B. subtilis sposoben sorodstvenega razlikovanja med genetsko sorodnimi in manj sorodnimi sevi ter z njimi vzpostavlja različne tipe interakcij. To je med drugim razvidno ob stiku dveh rojev, kjer se med manj sorodnimi sevi zaradi antagonističnih odnosov oblikujejo jasno izražene meje. Namen magistrske naloge je bil raziskati vpliv sorodstvenega razlikovanja na prenos DNA med genetsko sorodnimi in manj sorodnimi sevi ter med različnimi vrstami rodu Bacillus v tekočem in na poltrdnem gojišču. Predvidevali smo, da se bo učinkovitost prenosa in vgraditve DNA zmanjševala z upadanjem genetske podobnosti med donorskim in recipientskim sevom. V ta namen smo pripravili 43 mutantnih sevov rodu Bacillus odpornih proti rifampicinu, ki so služili kot donorski sevi za prenos DNA v recipientski sev B. subtilis PS-216. V okviru naloge smo vpeljali in optimizirali metodo za izolacijo mutant odpornih proti rifampicinu ter identificirali točkovne mutacije v genu rpoB. Ugotovili smo, da v tekočem gojišču prihaja do spontanih mutant na rifampicin, zato vpliva sorodnosti DNA na transformacijo recipientskega seva nismo mogli uspešno določiti. Preverjali smo tudi prenos DNA na poltrdnem gojišču in pokazali trend upadanja frekvence prenosa DNA z manjšanjem stopnje sorodnosti med donorskim in recipientskim sevom. Ugotovili smo, da je vgraditev še mogoča pri 84,90 % podobnosti na podlagi primerjave zaporedja gena rpoB med recipientskim in donorskim sevom. Z nalogo smo nakazali, da genetska sorodnost vpliva na frekvenco prenosa in vgraditve DNA med donorskim in recipientskim sevom ter s tem poglobili vpogled v pretok genov med različnimi sevi in vrstami rodu Bacillus. Bacillus subtilis is a naturally competent bacterium capable of taking up extracellular DNA from the environment and integrating it into its chromosome. The efficiency of this process strongly depends on the degree of genetic relatedness between donor DNA and the recipient genome and decreases markedly as genetic similarity declines. In addition, B. subtilis exhibits kin discrimination, forming different interactions with genetically related and less related strains, which is evident at swarm boundaries where antagonistic interactions lead to the formation of clear demarcation lines. The aim of this master’s thesis was to investigate the impact of kin discrimination on DNA transfer between genetically related and less related strains, as well as between different species of the genus Bacillus, in liquid and semi-solid media. We hypothesized that DNA transfer and integration efficiency would decrease with declining genetic similarity between donor and recipient strains. To address this, we generated 43 rifampicin-resistant Bacillus mutant strains as donors for DNA transfer to the recipient strain B. subtilis PS-216, optimized a method for isolating rifampicin-resistant mutants, and identified point mutations in the rpoB gene. Spontaneous rifampicin-resistant mutants arose in liquid medium, preventing reliable assessment of the effect of DNA relatedness on transformation under these conditions. In contrast, experiments on semi-solid medium revealed a trend in decreasing DNA transfer frequency with decreasing donor–recipient relatedness. DNA integration remained detectable at 84,90 % sequence similarity based on rpoB gene comparisons between the recipient strain and donor strain PS-99. Overall, these results indicate that genetic relatedness significantly influences DNA transfer and integration, providing insight into gene flow among different Bacillus strains and species. Bacillus subtilis odpornost proti rifampicinu gen rpoB frekvenca prenosa DNA sorodnost Bacillus subtilis rifampicin resistance rpoB gene DNA exchange frequency relatedness true false false [Ž. Guček] Slovenski jezik Angleški jezik Magistrsko delo/naloga 2026-03-19 07:15:17 2026-03-19 07:15:30 2026-03-21 04:07:18 0000-00-00 00:00:00 2026 0 0 0000-00-00 NiDoloceno NiDoloceno NiDoloceno 0000-00-00 0000-00-00 0000-00-00 579.25:579.852.11 279144 272285187 56926.pdf 56926.pdf 1 3F6B314A8E8845B3F454BD2DA9B6DC57 1d7a10571677a96e2c2e2e0fa7e215999402434176e12b1d2d8bffce05051d2d 035279e7-235b-11f1-b0ab-0050569b8976 https://repozitorij.uni-lj.si/Dokument.php?lang=slv&id=230829 Biotehniška fakulteta 0 0 0