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Plasmid-encoded MCP is involved in virulence, motility, and biofilm formation of Cronobacter sakazakii ATCC 29544 : 크로노박터 사카자키의 새로운 병원성 플라스미드 분리와 그 병원성 유전자 MCP의 특성 연구
| DC Field | Value | Language |
|---|---|---|
| dc.contributor.advisor | 유상렬 | - |
| dc.contributor.author | 황혜련 | - |
| dc.date.accessioned | 2017-07-14T06:41:30Z | - |
| dc.date.available | 2017-07-14T06:41:30Z | - |
| dc.date.issued | 2014-02 | - |
| dc.identifier.other | 000000018006 | - |
| dc.identifier.uri | https://hdl.handle.net/10371/125846 | - |
| dc.description | 학위논문 (석사)-- 서울대학교 대학원 : 농생명공학부, 2014. 2. 유상렬. | - |
| dc.description.abstract | The aim of this study was to elucidate the function of the plasmid-encoded mcp (methyl-accepting chemotaxis protein) gene that plays pleiotropic roles in C. sakazakii ATCC 29544. By searching for virulence factors using a random transposon insertion mutant library, I identified and sequenced a new plasmid, pCSA2, in C. sakazakii ATCC 29544. In silico analysis of pCSA2 revealed that it encoded 6 open reading frames, and one of them was mcp. The mcp mutant was defective for invasion into and adhesion to epithelial cells, and the virulence of the mcp mutant was attenuated in rat pups. In addition, I showed that putative MCP regulates the motility of C. sakazakii ATCC 29544 and the expression of the flagellar genes was enhanced in the absence of a functional mcp gene. Furthermore, lack of the mcp gene also impaired the ability of C. sakazakii to form biofilm. Our results demonstrate a regulatory role for MCP in diverse biological processes including virulence of C. sakazakii ATCC 29544. To the best of our knowledge, this study is the first to elucidate a potential function of a plasmid-encoded MCP homolog in C. sakazakii ST8. | - |
| dc.description.tableofcontents | ABSTRACT................................................................................................................i
CONTENTS.............................................................................................................iii I. INTRODUCTION...............................................................................................1 Ⅱ. MATERIALS AND METHODS........................................................................4 1. Bacterial strains, plasmids, and growth conditions.....................................4 2. Random mutagenesis and screening .............................................................4 3. Determination of the transposon insertion site …........................................5 4. Complete nucleotide sequencing and bioinformatics ……………………..5 5. Nucleotide sequence accession number ………………….………...……....6 6. Cell culture………...........................................................................................6 7. Site-specific mutagenesis of C. sakazakii ATCC29544................................7 8. Construction of the complementation strain ………………….…………..8 9. Invasion assay …………………….................................................................8 10. Adhesion assay.……......................................................................................9 11. In vivo rat pup virulence assay by competitive index analysis…………10 12. Motility assay.…………………………………....……………..…..…..…11 13. RNA isolation and qRT-PCR.……………………………………………12 14. Biofilm assay………………………………………………………………12 15. Two-dimensional gel electrophoresis (2DGE) …………………..………12 16. Statistical analysis.......................................................................................13 Ⅲ. RESULTS...........................................................................................................15 1. A plasmid-encoded gene discovered in an invasion-attenuated mutant ………………..…………………………………...…………………….……...15 2. pCSA2 is a novel plasmid in C. sakazakii ATCC 29544 that contains 6 open reading frames…………………………………………………………..18 3. Mcp protein is comprised of two sensory PAS domain and MCP domain................................................................................................................19 4. Plasmid-encoded putative MCP affects adhesion/invasion……...............21 5. C. sakazakii lacking a plasmid-encoded putative MCP showed attenuated virulence in rat pups……...............................................................…………..25 6. Plasmid-born MCP regulates motility of C. sakazakii ATCC 29544……27 7. Biofilm formation is affected by the plasmid-encoded putative MCP.....32 8. Lack of the mcp gene affected the expression of multiple genes in C. sakazakii..............................................................................................................34 9. MglB mutant has similar effect with mcp mutant..............................36 Ⅳ. DISSCUSSION...............................................................................................38 Ⅴ. REFFERENCES....................................................................................41 국문초록.......................................................................................................51 | - |
| dc.format | application/pdf | - |
| dc.format.extent | 914623 bytes | - |
| dc.format.medium | application/pdf | - |
| dc.language.iso | en | - |
| dc.publisher | 서울대학교 대학원 | - |
| dc.subject | Cronobacter sakazakii | - |
| dc.subject | virulence | - |
| dc.subject | methyl-accepting chemotaxis protein | - |
| dc.subject | pCSA2 | - |
| dc.subject | D-galactose-binding periplasmic protein | - |
| dc.subject.ddc | 630 | - |
| dc.title | Plasmid-encoded MCP is involved in virulence, motility, and biofilm formation of Cronobacter sakazakii ATCC 29544 | - |
| dc.title.alternative | 크로노박터 사카자키의 새로운 병원성 플라스미드 분리와 그 병원성 유전자 MCP의 특성 연구 | - |
| dc.type | Thesis | - |
| dc.description.degree | Master | - |
| dc.citation.pages | 55 | - |
| dc.contributor.affiliation | 농업생명과학대학 농생명공학부 | - |
| dc.date.awarded | 2014-02 | - |
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