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Vloga genskih skupkov, sestavljenih iz genov podobnih susCD in genov kodirajočih za nespecifične nukleaze, v razgradnji in uporabi eksogenih nukleinskih kislin pri rodovih Prevotella in Bacteroides
Pinterič, Bor (Author), Accetto, Tomaž (Mentor) More about this mentor... This link opens in a new window

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Abstract
Namen magistrskega dela je bil raziskati vlogo genskih skupkov, sestavljenih iz genov podobnih susCD in genov, kodirajočih za nespecifične nukleaze. Osredotočili smo se na vrste iz rodov Prevotella in Bacteroides in njihove sposobnosti privzema in razgradnje različnih polisaharidov s pomočjo genskih skupkov sestavljenih iz genov za privzem in razgradnjo različnih polisaharidov (PUL-i). Obravnavali smo varianto skupkov, katerih tarčne molekule so predvidoma nukleinske kisline, gene za razgradnjo saharidov pa nadomestijo nespecifične nukleaze. Delo je obsegalo tri glavne sklope: bioinformacijski pregled, gojitvene študije in opredelitev aktivnosti skupkov z RT-qPCR. V genomih smo iskali prisotnost podobnih skupkov. Na šestih izbranih sevih smo s pomočjo Hungate gojitvenih tehnik pokazali sposobnost rasti v gojišču z dodatkom posameznih nukleozidov, sladkorjev (riboze oz. deoksiriboze) ter DNA ali RNA. Izražanje genov iz domnevno aktivnih skupkov smo potrdili z uporabno reverzne transkripcije izolirane RNA, ter PCR v realnem času. Izkaže se, da so skupki široko zastopani, arhitekturno ohranjeni, ter da sevi kažejo sposobnost rasti v prisotnosti (gradnikov) nukleinskih kislin. Razlike v izražanju genov so opazne, a jih nismo uspeli točno razložiti, smo pa nakazali morebiten pomen skupkov v razgradnji nukleinskih kislin, tako pri vampnih simbiontih, kot tudi potencialnih patogenih predstavnikih ustne mikrobiote.

Language:Slovenian
Keywords:molekularna genetika, genski skupki, susC, susD, nucA, nucB, nespecifične nukleaze, Prevotella, Bacteroides, zunajcelične nukleinske kisline, razgradnja, uporaba, anaerobne tehnike, gelska elektroforeza, reverzna transkripcija, PCR v realnem času
Work type:Master's thesis/paper (mb22)
Tipology:2.09 - Master's Thesis
Organization:BF - Biotechnical Faculty
Year:2019
Publisher:[B. Pinterič]
UDC:601.4:577.21:616-093
COBISS.SI-ID:5137784 Link is opened in a new window
Views:260
Downloads:151
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Secondary language

Language:English
Title:The role of gene clusters, composed of genes similar to susCD and those coding for nonspecific nucleases, in the degradation and utilization of exogenous nucleic acids among Prevotella and Bacteroides
Abstract:
The purpose of this master's thesis was to determine the role of gene clusters, comprised of genes similar to susCD and genes coding for non-specific nucelases. The work was done on species of the genera Prevotella and Bacteroides, known for their ability to degrade and utilize various polysaccharides, which is most commonly achieved via PULs. Our work was focused on a variant of these clusters presumably targeting nucleic acids and the genes for saccharide degradation are replaced by nucleases. The work encompassed three major parts: bioinformatical overview, growth studies and RT-PCR to assess cluster activity. We searched for the presence of such clusters in the genomes of the Prevotella and Bacteroides genera. Using the Hungate technique we showed the ability of six chosen strains to grow in media with the addition of specific nucleosides, sugars (ribose, deoxyribose) and DNA or RNA. Expression of the putatively active clusters was monitored with by reverse transcription of the isolated RNA and real-time PCR. The clusters appear to be widely present as well as architecturally conserved and the tested strains have been able to grow on select media. Differences in the gene expression levels are noticable, but despite being unable to explain them completely, these conclusions show that the clusters might play an important role in both symbiotic rumen strains and potential pathogens from the oral cavity.

Keywords:molecular genetics, gene clusters, susC, susD, nucA, nucB, non-specific nucleases, Prevotella, Bacteroides, extracellular nucleic acids, degradation, utilization, anaerobic techniques, gel electrophoresis, reverse transcription, real-time PCR

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