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完整後設資料紀錄
DC 欄位 | 值 | 語言 |
---|---|---|
dc.contributor.advisor | 柯淳涵(Chun-Han Ko) | |
dc.contributor.author | Heng-Yi Li | en |
dc.contributor.author | 李姮誼 | zh_TW |
dc.date.accessioned | 2021-06-13T01:03:04Z | - |
dc.date.available | 2017-12-31 | |
dc.date.copyright | 2007-07-28 | |
dc.date.issued | 2007 | |
dc.date.submitted | 2007-07-25 | |
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dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/29228 | - |
dc.description.abstract | 木聚醣酶(Xylanase)在工業上具有相當廣泛的應用價值,它在製漿造紙業、食品飲料工業、飼料業、紡織業皆有其應用。本實驗室之前已將一段由台紙洗漿黑液中分離之Paenibacillus campinanesis BL11的木聚醣酶基因轉殖入大腸桿菌中,並利用 lac 啟動子來控制基因的表現,可以有效的在宿主細胞體內合成木聚醣酶,選殖出的木聚醣酶基因是由1,131個核苷酸所組成,經由轉錄及轉譯可得到大小為41kDa的木聚醣酶。本研究乃探討此重組大腸桿菌之木聚醣酶最適生產條件,並建立一套控制的饋料批次醱酵系統,藉由饋料批次培養更進一步提升木聚醣酶的生產效率。在搖瓶培養規模中所測試的項目為宿主種類、培養基之碳源濃度、有機氮源種類、無機氮源種類。在以搖瓶所測試出之最適合培養基作為五公升醱酵槽規模中起始培養基組成份,其主要組成份為0.5%葡萄糖、0.5%蛋白腖、0.2% K2HPO4、0.3% KH2PO4、0.5% (NH4)2HPO4、0.05% MgSO4•7H2O及微量元素。以E. coli BL21 (DE3)為宿主在控制條件為pH 7.0,溶氧值控制在35%,溫度恒定為37oC,於培養第6小時加入 0.06 mM IPTG誘導木聚醣酶表現,於培養30小時之菌體量可達 2.63 g DCW/L, 木聚醣酶比活性233.45IU/mg。以純化後的木聚醣酶水解樺木及燕麥木聚醣的產物以木三糖為主。 | zh_TW |
dc.description.abstract | Xylanase have rather extensive application value on the industry, it is making syrup deckle industry, the food beverage industry, the forage industry, textile industry all has it an application. This laboratory before already there is scholar will a by from Hsinying Paper Mill of Taiwan Pulp and Paper of the BL11 of Paenibacillus campinanesis of the separation in the black liquid to gather the xylanase gene turns to go into Escherichia coli, and makes use of the lac promoter to control the performance of the gene, can effectively of expression xylanase in the host cell body. The cloned BL11 xylanase is composed of 1,131 nucleotides and can encode a protein of 41 kDa. The effects of culture variables on xylanase production by the recombinant E. coli were investigated using a controlled fed-batch fermentation system. In flask culture scale test of the item be having host category, carbon source density, organic nitrogen source category of developing medium and having no machine nitrogen source category. With E. coli BL21 (DE3) for the host be controling the condition as pH 7.0, dissolve the oxygen value control in 35%, the temperature in 37 oC, joining the 0.06 mM IPTG inducement in culture 6 h to gather xylanase after culture 30h, can reach to 2.63 g DCW/L and xylanase specific activity reach to 233.45 IU/mg. Xylotriose was the major hydrolytic product of oat spelt and birchwood xylan. | en |
dc.description.provenance | Made available in DSpace on 2021-06-13T01:03:04Z (GMT). No. of bitstreams: 1 ntu-96-R94625027-1.pdf: 810488 bytes, checksum: 6d09093e8f5bc4e8a70f5f41985b761c (MD5) Previous issue date: 2007 | en |
dc.description.tableofcontents | Index i
Table index iv Figure index v 摘要 viii Abstract ix Chapter 1 Introduction 1 Chapter 2 Literature review 4 2.1 Character of Paenibacillus sp. isolate BL11 4 2.2 Application of xylanase 5 2.3 Factors that affect fermentation culture of E. coli 8 2.4 The control principle of the fed-batch culture 16 Chapter 3 Material and Methods 18 3.1 The structure of gene 18 3.2 Bacterial strain 20 3.3 Cell growth of shaking flask 21 3.4 The determination of plasmid stability 22 3.5 Xylanase production in flask culture 23 3.6 Cell density measurements 24 3.7 Polysaccharide hydrolase assay 25 3.7.1 Sodium dodecylsulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and zymography 25 3.7.2 DNSA (dinitrosalicylic acid) assay 26 3.8 Glucose and acetate concentration assay 27 3.9 Preparation of cell samples for protein product 28 3.10 Fermentation 28 3.10.1 The system of fermenter 28 3.10.2 The fermentation medium 29 3.10.3 Batch cultivation 30 3.10.4 Fed-batch cultivation 31 3.11 Purification of xylanase 32 3.11.1 Disruption of cells 32 3.11.2 Affinity chromatography 32 3.11.3 Protein concentration assay 33 3.12 Hydrolysis products of birchwood xylan and oat spelt xylan 34 Chapter 4 Results and Discussion 35 4.1 Cell growth of recombinant E. coli in shaking flask 35 4.2 The determination of plasmid stability 44 4.3 Xylanase production in flask culture 45 4.4 Preparation of cell samples for protein product 53 4.5 Fermentation 54 4.5.1 Batch fermentation 54 4.5.2 Fed-batch cultivation 56 4.6 Purification of xylanase 61 4.7 Hydrolysis products of birchwood xylan and oat spelt xylan 63 Chapter 5 Conclusion 68 Chapter 6 References 69 | |
dc.language.iso | en | |
dc.title | 以大腸桿菌饋料批次生產小枯草桿菌BL11之熱穩定重組木聚醣酶 | zh_TW |
dc.title | Fed-Batch Production of Thermostable Recombinant Xylanases from Paenibacillus sp. isolate BL11 in Escherichia coli | en |
dc.type | Thesis | |
dc.date.schoolyear | 95-2 | |
dc.description.degree | 碩士 | |
dc.contributor.oralexamcommittee | 劉佳振(Chia-Chen Liou),張上鎮(Shang-Tzen Chang),杜鎮(Jenn Tu),蕭英倫(Ying-Lun Hsiao) | |
dc.subject.keyword | 木聚醣酶,重組大腸桿菌,饋料批次發酵, | zh_TW |
dc.subject.keyword | Xylanase,Recombinant E. coli,Fed-batch fermentation., | en |
dc.relation.page | 74 | |
dc.rights.note | 有償授權 | |
dc.date.accepted | 2007-07-25 | |
dc.contributor.author-college | 生物資源暨農學院 | zh_TW |
dc.contributor.author-dept | 森林環境暨資源學研究所 | zh_TW |
顯示於系所單位: | 森林環境暨資源學系 |
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