DC 欄位 | 值 | 語言 |
dc.contributor | 鄭秋萍 | zh-TW |
dc.contributor | Cheng, Chiu-Ping | en |
dc.contributor | 臺灣大學:植物科學研究所 | zh-TW |
dc.contributor.author | 王冠中 | zh-TW |
dc.contributor.author | Wang, Kuan-Chung | en |
dc.creator | 王冠中 | zh-TW |
dc.creator | Wang, Kuan-Chung | en |
dc.date | 2009 | en |
dc.date.accessioned | 2010-05-11T01:04:17Z | - |
dc.date.accessioned | 2018-07-06T03:41:46Z | - |
dc.date.available | 2010-05-11T01:04:17Z | - |
dc.date.available | 2018-07-06T03:41:46Z | - |
dc.date.issued | 2009 | - |
dc.identifier.other | U0001-0508200916165400 | en |
dc.identifier.uri | http://ntur.lib.ntu.edu.tw//handle/246246/181955 | - |
dc.description.abstract | 青枯菌為土壤中常見的植物病原菌之一,可感染超過兩百種植物,其中包含番茄、馬鈴薯等極具經濟價值的作物。土壤中的青枯菌侵入植物體後,可在木質部大量增殖並分泌胞外多醣或是細胞壁分解酵素,受到感染的植物常因為木質部水分運輸受阻進而造成全株死亡。針對青枯菌造成的細菌性萎凋病,到目前為止尚無有效的藥劑可以預防或是控制其病徵產生。因此生物性防治法便成為一個值得期待的防治方法。為了瞭解青枯病菌與其噬菌體間之交互作用,並期許可以應用於青枯病的防治,本論文初步在台灣數個地區的土壤中分離青枯菌的噬菌體。由基因組隨機定序結果得知本研究目前所分離之噬菌體與T7類有高度的胺基酸相似性,並且對其進行了基本的特性分析、宿主範圍測試和溶菌效果分析等,而進一步的生物防治測試也證明外加噬菌體確實可降低番茄青枯病的發生率。此外,為了更加了解噬菌體與青枯菌的分子層級交互關係,本研究大量篩選青枯菌跳躍子插入之突變株庫,發現二十一個與噬菌體感染相關之青枯菌基因;其中約半數是屬於表面脂多糖 (lipopolysaccharides) 生合成相關酵素基因,其餘則為與ABC類運送蛋白或甘露糖運輸與代謝相關基因,以及某些尚待研究的新穎基因。透過這些分子層面的研究使得我們對青枯菌及其噬菌體交互關係有更進一步了解,且對研擬病害生物防治策略也提供了許多重要的啟發。 | zh-TW |
dc.description.abstract | Ralstonia solanacearum is a soil-borne, xylem-inhabited plant pathogen. The wide host range (Hayward, 1991) and the lethal wilting symptom render R. solanacearum a globly important pathogen. Commercial pesticides and antibiotics are generally ineffective in controlling bacterial wilt (BW) caused by the pathogen (Denny, 2006). Therefore, biological control methods would be one of the feasible approaches to reduce crop losses caused by this bacterium. In this study, indigenous R. solanacearum bacteriophages were isolated and identified to be T7-like phages. The characteristics of these phages, including host range, lytic ability and optimal pH for the isolated phages have been determined. Their potential of being employed as biocontrol agents against tomato BW was also demonstrated. Furthermore, a systematic survey of R. solanacearum genes essential for phage infection has been carried out and led to the identification of 21 genes. About a half of these genes are or could be involved in biosynthesis of lipopolysaccharides (LPS). The rest of the identified genes included those involved in sugar metabolism and unknown function genes. The elucidation of phage infection mechanism not only is valuable for basic research, but also could be useful for boosting the efficiency of phage therapy by artificially introducing infection essential genes into the phage genomes. These studies collectively are expected to pave the way for elucidating phage-host interactions and establishing useful biocontrol means. | en |
dc.description.tableofcontents | Abbreviation i文摘要 iibstract iiiontents ivhapter 1 solation and characterization of Ralstonia solanacearum phages and assessment of phage biocontrol strategies 1ntroduction 1ature of bacteriophages 1oles of bacteriophages in ecology and evolution with bacteria 2iocontrol potential of bacteriophages 3actors affecting the efficiency of phage-based biocontrol means 4nhancement of phage biocontrol efficiency 5alstonia solanacearum and bacterial wilt 6he studies of R. solanacearum phages 7iological-based control approaches against BW 7otivation of the study 8aterials and methods 9acteria strain and culture condition 9acteriophage isolation and enrichment 10ost range determination 11hage DNA isolation and characterization 12hage lysis assay 13hage φ29 purification and concentration 13ransimission electronic microscopic 14tructural protein analysis of phage φ29 by SDS-PAGE 14hermal inactivation and pH tolerance assay of φ29 15iocontrol and tomato growth condition 15ioinformatics analysis tools in the study 16esults 17solation of novel T7-like bacteriophage of R. solanacearum in Taiwan. 17etermination of host range 17andom sequencing revealed unique T7-like of phages in our study 18nhibition of R. solanacearum growth in tube by phages 19EM morphology revealed T7-like phage morphology of phage φ29 20he structural proteins of φ29 were different to known T7-like phage 20hage φ29 was rapidly inactivated at temperature above 60℃ 21hage φ29 is stable at pH 6-8 21upplements ofφ29 in soils reduced the likelihood of BW 21pplication of hrpG mutants didn’t improve the biocontrol effect 22iscussion 24solation of novel T7-like R. solanacearum phages in soil in Taiwan 2429- similar sequence was existed in other prokaryotes as putative prophage sequence 2429- similar sequence was found in Populus trichocarpa (Black cottonwood) 25ost range of the isolated phages R. solanacearum 26emperature and pH tolerance of φ29 27he ratio of inoculated bacteriophage affect the biocontrol efficiency 27he inoculating timing of bacteriophage affect the biocontrol efficiency 28upplement of hrpG mutant did not lead to enhance biocontrol efficiency 29ariation of plant growth condition of affected biocontrol efficiency 30uggested future work 31hapter 2 enome-wide survey of R. solanacearum genes involved in phage infection by screening transposon mutagenesis library 34ntroduction 347-like bacteriophages and their life cycles 34acterial receptor for phage infection 35tudies of bacterial genes essential for phage growth 36hage modification and application approaches 37otivation of our study 37aterials and Methods 39acteria strain and culture condition 39onstruction of R. solanacearum transposon-insertion mutant library 39creening phage-resistant mutants and EOP assay 40dentification and confirmation of disruptive genes. 41ntibiotic stress response assay of mutants 43ioinformatics analysis tools in the study. 44esults 45creening for R. solanacearum phage-resistant mutants 45dentification of disruptive loci in R. solanacearum mutants 45etermation of the efficiency of plaquing (EOP) of the phage-resistant mutants 46rowth of the R. solanacearum mutants on different media. 47rowth of R. solanacearum mutants in the presence of SDS. 47rowth of R. solanacearum mutants on SM1 medium. 48ensitivity of R. solanacearum mutants to antibiotics. 48iscussion 50ipopolysacchrides core biosynthesis 50ipopolysacchrides O antigen biosynthesis 53utative cell wall biogenesis 56annose metabolism related 58BC-type transporters-like proteins and vacJ 60ovel genes 63ossible function of phage-growth essential genes in φ29 life cycle 64ffectiveness and other concerns of the mutant screening 66uggested future work 67eferences 68ables 78able 1. Host range of R. solanacearum phages identified in our study 78able 2. random sequencing of phage genome digestion fragments 80able 3. The wilting symptom on tomato cv. L390 infected with R. solanacearum strain Pss4 (different concentration) in response to application of phage at different timing 82able 4. The wilting symptom on tomato cv. L390 infected with R. solanacearum strain Pss4 in response to application of phage and hrpG mutants at different timing 83able.5 In vitro growth and EOP of R. solanacearum mutants 84able 6. Phage growth essential genes in R .solanacearum identified in this study 85igures 86igure 1. Host range of φ29 by spot test. 86igure 2. blastx dot matrix views of alignment of φ29 fragments and φRSB1 87igure 3: Control of growth of R. solanacearum Pss4 in the presence of phages 88igure 4. Electron micrographs of R. solanacearum φ29 89igure 5. Structural proteins of φ29 by SDS-PAGE separation 90igure 6. Thermal inactivation regression line of phage φ29 at 60℃. 91igure 7. pH tolerance of phage φ29. 92igure 8. Progress of disease severity on tomato cultivar L390 in response to R. solanacearum and/ or phage φ29 inoculation 93igure 9. Progress of disease severity on tomato cultivar L390 in response to R. solanacearum and/ or phage φ29 and hrpG mutant inoculation 94igure 10. Relatively growth condition of phage resistant mutants in response 95igure 11. Genomic localization of Tn5 insertion in phage resistance mutants. 99upplementary tables 100able S1. Primers used in the study 100able S2. Locations of soil samples isolated in Taiwan 102able S3. TAIL-PCR program in this study 103able S4. TAIL-PCR recipes in this study 104able S5. Summary of screening results in this study 105upplementary figures 106igure S1. Digestion pattern of isolated phage gDNA 107igure S2. Structural protein banding patterns of other similar T7-like phages 108igure S3. pCRII TOPO vector map 109igure S4. EZ-Tn5 <KAN-2> transposon map 110igure S5. Ralstonia solanacearum LPS biosynthesis pathway 111igure S6. Identified genes involved in LPS biosynthesis pathway 112 | en |
dc.format | application/pdf | en |
dc.format.extent | 3718969 bytes | - |
dc.format.mimetype | application/pdf | - |
dc.language | en | en |
dc.language.iso | en_US | - |
dc.subject | 青枯菌 | zh-TW |
dc.subject | 細菌性萎凋 | zh-TW |
dc.subject | 噬菌體 | zh-TW |
dc.subject | 生物防治 | zh-TW |
dc.subject | 脂多醣 | zh-TW |
dc.subject | Ralstonia | en |
dc.subject | bacterial wilt | en |
dc.subject | bacteriophage | en |
dc.subject | biocontrol | en |
dc.subject | lipopolysaccharides | en |
dc.title | 植物青枯病菌及其噬菌體交互關係之探究 | zh-TW |
dc.title | Elucidation of the interaction between Ralstonia solanacearum and its bacteriophages | en |
dc.identifier.uri.fulltext | http://ntur.lib.ntu.edu.tw/bitstream/246246/181955/1/ntu-98-R96b42004-1.pdf | - |
item.fulltext | with fulltext | - |
item.grantfulltext | open | - |
item.languageiso639-1 | en_US | - |
顯示於: | 植物科學研究所
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