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完整後設資料紀錄
DC 欄位 | 值 | 語言 |
---|---|---|
dc.contributor.advisor | 王愛玉(Ai-Yu Wang) | |
dc.contributor.author | Jiun-Wei Fan | en |
dc.contributor.author | 范峻維 | zh_TW |
dc.date.accessioned | 2021-06-08T04:38:57Z | - |
dc.date.copyright | 2009-08-19 | |
dc.date.issued | 2009 | |
dc.date.submitted | 2009-08-17 | |
dc.identifier.citation | References:
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Molecular cloning, expression and characterization of a novel apoplastic invertase inhibitor from tomato (Solanum lycopersicum) and its use to purify a vacuolar invertase. Biochimie 90(11-12):1611-1623. Roitsch T, Gonzalez MC. 2004. Function and regulation of plant invertases: sweet sensations. Trends Plant Sci 9(12):606-613. Rosenkranz H, Vogel R, Greiner S, Rausch T. 2001. In wounded sugar beet (Beta vulgaris L.) tap-root, hexose accumulation correlates with the induction of a vacuolar invertase isoform. J Exp Bot 52(365):2381-2385. Sander A, Krausgrill S, Greiner S, Weil M, Rausch T. 1996. Sucrose protects cell wall invertase but not vacuolar invertase against proteinaceous inhibitors. FEBS Lett 385(3):171-175. Schwimmer S, Makower RU, Rorem ES. 1961. Invertase & invertase inhibitor in potato. Plant Physiol 36(3):313-316. Sergeeva LI, Keurentjes JJ, Bentsink L, Vonk J, van der Plas LH, Koornneef M, Vreugdenhil D. 2006. Vacuolar invertase regulates elongation of Arabidopsis thaliana roots as revealed by QTL and mutant analysis. Proc Natl Acad Sci U S A 103(8):2994-2999. Sherson SM, Alford HL, Forbes SM, Wallace G, Smith SM. 2003. Roles of cell-wall invertases and monosaccharide transporters in the growth and development of Arabidopsis. J Exp Bot 54(382):525-531. Silke K, Steffen G, Ulrike K, Rolf V, Thomas R. 1998. <b>In transformed tobacco cells the apoplasmic invertase inhibitor operates as a regulatory switch of cell wall invertase</b>. The Plant Journal 13(2):275-280. Swarbrick PJ, Schulze-Lefert P, Scholes JD. 2006. Metabolic consequences of susceptibility and resistance (race-specific and broad-spectrum) in barley leaves challenged with powdery mildew. Plant Cell Environ 29(6):1061-1076. Tang GQ, Luscher M, Sturm A. 1999. Antisense repression of vacuolar and cell wall invertase in transgenic carrot alters early plant development and sucrose partitioning. Plant Cell 11(2):177-189. Vargas WA, Pontis HG, Salerno GL. 2008. New insights on sucrose metabolism: evidence for an active A/N-Inv in chloroplasts uncovers a novel component of the intracellular carbon trafficking. Planta 227(4):795-807. Voegele RT, Wirsel S, Moll U, Lechner M, Mendgen K. 2006. Cloning and characterization of a novel invertase from the obligate biotroph Uromyces fabae and analysis of expression patterns of host and pathogen invertases in the course of infection. Mol Plant Microbe Interact 19(6):625-634. von Schaewen A, Stitt M, Schmidt R, Sonnewald U, Willmitzer L. 1990. Expression of a yeast-derived invertase in the cell wall of tobacco and Arabidopsis plants leads to accumulation of carbohydrate and inhibition of photosynthesis and strongly influences growth and phenotype of transgenic tobacco plants. EMBO J 9(10):3033-3044. Weil M, Krausgrill S, Schuster A, Rausch T. 1994. A 17-kDa Nicotiana tabacum cell-wall peptide acts as an in-vitro inhibitor of the cell-wall isoform of acid invertase. Planta 193(3):438-445. Yau YY, Simon PW. 2003. A 2.5-kb insert eliminates acid soluble invertase isozyme II transcript in carrot (Daucus carota L.) roots, causing high sucrose accumulation. Plant Mol Biol 53(1-2):151-162. 鄭瑞芬 (1985) 綠竹筍蔗糖轉化酶進一步的純化研究。碩士論文,國立臺灣大學農業化學研究所。 莊榮輝 (1985) 水稻蔗糖合成酶之研究。博士論文,國立臺灣大學農業化學研究所。 劉力閣 (2002) 綠竹筍蔗糖轉化酶 cDNA 之選殖與鑑定。碩士論文,國立臺灣大學農業化學研究所。 王俐婷 (2003) 甘藷葉酸性蔗糖轉化酶之分子與生化檢定。博士論文,國立臺灣大學農業化學研究所。 王永樑 (2003) 甘藷葉可溶性酸性蔗糖轉化酶及其抑制因子之研究。博士論文,國立臺灣大學農業化學研究所。 謝常文 (2004) 綠竹筍蔗糖轉化酶在酵母菌 Pichia pastoris 中的表現與檢定。碩士論文,國立臺灣大學微生物與生化學研究所。 陳泰宏 (2006) 以定位點突變法探討綠竹液泡型蔗糖轉化酶活性區之結構與功能。碩士論文,國立臺灣大學微生物與生化學研究所。 | |
dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/23039 | - |
dc.description.abstract | 蔗糖轉化酶 (EC 3.2.1.26, β-D-fructofuranosidase),為高等植物中參與蔗糖代謝其中一個重要酵素,可水解蔗糖產生葡萄糖及果糖。蔗糖轉化酶對於植物生長及發育扮演重要的角色。綠竹筍 (Bambusa oldhamii) 中至少擁有三種不同型態的蔗糖轉化酶,過去研究也指出綠竹筍中可能有蔗糖轉化酶活性的調控因子存在。本研究利用酵母菌 Picahia pastoris 表現重組蛋白質 rBoIT2 和 rBoIT3 作為分析綠竹筍中蔗糖轉化酶活化因子活性測定所使用。
綠竹筍的可溶性蛋白質粗抽液經硫酸銨分劃後發現,酸性蔗糖轉化酶在飽和濃度 50 ~ 80% 的硫酸銨分劃中具有最高的比活性;而在飽和濃度 50 ~ 100% 的分劃中含有可提升 rBoIT2 活性的活化因子。硫酸銨分劃經熱處理後內生性蔗糖轉化酶活性會被除去,但活化因子具有很高的熱穩定性。活化因子利用 Sepharose CL-6B 膠體過濾層析再經 Mono Q 陰離子交換層析純化,或是先進行DEAE-Sephacel 陰離子交換層析再經 Superdex G200 膠體過濾層析純化。這兩種純化方法的結果以 SDS-PAGE 檢定呈現出相似的蛋白質色帶。在分子量大小 60 kD, 45 kD, 38 kD, 25 kD 可以觀察到蛋白質色帶。純化結果顯示綠竹筍中極有可能存在蔗糖轉化酶的活化因子 | zh_TW |
dc.description.abstract | Invertase (EC 3.2.1.26, β-D-fructofuranosidase) is one of the major enzymes involved in sucrose metabolism of higher plants. The enzyme catalyzes the hydrolysis of sucrose to glucose and fructose, and is crucial for plant growth and development. There are at least three invertase isozymes in green bamboo (Bambusa oldhamii), and inhibitor and activator of bamboo invertase have been found in previous studies. To purify and characterize the activator of invertase from bamboo, the recombinant proteins, rBoIT2 and rBoIT3 were purified from Picahia pastoris for analysis of the activities of activator.
Soluble proteins from green bamboo shoots were fractionated by ammonium sulfate. The highest specific activity of invertase was found in the fraction of 50 ~ 80% saturation of ammonium sulfate. Activators were present in the fraction of 50 ~ 100% saturation of ammonium sulfate. After heat treatment, the activities of invertase were removed, while activators showed high thermo stability. Activators were purified by Sepharose CL-6B gel-filtration chromatography and then by Mono Q ion-exchange chromatography or first by DEAE-Sephacel ion-exchange chromatography and then by Superdex G200 gel-filtration chromatography. SDS-PAGE analysis of proteins purified by these two methods showed a similar protein pattern, which contained four major proteins with molecular masses of 60 kD, 45 kD, 38 kD, 25 kD, respectively. The results of this study indicated that the activator of invertase did exist in green bamboo. | en |
dc.description.provenance | Made available in DSpace on 2021-06-08T04:38:57Z (GMT). No. of bitstreams: 1 ntu-98-R96b47214-1.pdf: 1366093 bytes, checksum: 4b0096ec4e5d104967080add73aa7544 (MD5) Previous issue date: 2009 | en |
dc.description.tableofcontents | 目錄.....I
縮寫表...IV 摘要.....V ABSTRACT.VI 第一章 緒論..................................1 第一節 蔗糖與蔗糖降解酵素....................1 第二節 蔗糖轉化酶的酵素研究..................1 2.1 蔗糖轉化酶 1 2.2 蔗糖轉化酶的分類 2 2.2.1 酸性蔗糖轉化酶 2 2.2.2 中/鹼性蔗糖轉化酶 3 第三節 蔗糖轉化酶的生理角色..................4 3.1 蔗糖轉化酶參與醣類的分配 4 3.2 蔗糖轉化酶參與植物的發育 5 3.3 蔗糖轉化酶參與逆境反應 5 第四節 蔗糖轉化酶活性調節因子................6 4.1 蔗糖轉化酶的抑制因子 6 4.1.1 抑制因子分類與結構 6 4.1.2 抑制因子在植物體的生理功能 7 4.2 抑制因子的應用 7 4.3 蔗糖轉化酶的活化因子 8 第五節 實驗室在蔗糖轉化酶的研究基礎..........8 第六節 研究目的及實驗方向....................9 第二章 材料與方法..............................10 第一節 實驗材料..............................10 1.1 表現菌株 10 1.2 綠竹筍 10 第二節 實驗藥品..............................10 2.1 一般化學藥品 10 2.2 培養基 10 第三節 實驗儀器..............................11 3.1 離心機 11 3.2 電白質電泳、轉印系統 11 3.3 震盪培養箱 11 3.4 其他 11 第四節 實驗方法..............................12 4.1 重組蛋白質的表現 12 4.2 蔗糖轉化酶活化因子的純化 13 4.2.1 綠竹筍蛋白質粗萃取 13 4.2.2 可溶性蛋白質中純化活化因子 14 4.2.2.1 膠體過濾法 (gel-filtration) 14 4.2.2.2 離子交換法 (ion-exchange) 14 4.2.3 細胞壁蛋白質的萃取 15 4.3 蛋白質定量法 17 4.4 蔗糖轉化酶的活性測定 17 4.4.1 Dinitrosalicylic acid assay 17 4.4.2 Somogyi-Nelson’s method 18 4.5 蔗糖轉化酶調控因子之活性測定 19 4.6 聚丙醯胺膠體電泳 19 4.7 膠體染色法 20 4.7.1 Coomassie brilliant blue R-250 (CBR) 染色法 20 4.7.2 硝酸銀染色法 20 4.7.3 負染色法 (negative staining) 21 4.8 西方轉印法 21 4.9 免疫染色法 22 4.9.1 Diaminobenzidine (DAB) 呈色法 22 4.9.2 化學冷光呈色法 23 4.10 免疫共沉澱法 23 4.10.1 配製 Protein A Sepharose 膠體溶液 23 4.10.2 免疫共沉澱反應 23 第三章 結果與討論............................25 第一節 於酵母菌 (PICHIA PASTORIS) 中表現重組蛋白質25 第二節 硫酸銨分劃與熱處理條件探討. ..........26 2.1 可溶性蛋白質硫酸銨分劃 26 2.2 熱處理條件 27 第三節 從可溶性蛋白質中純化活化因子..........28 第四節 細胞壁蛋白質中調節因子之純化..........32 第五節 探討蔗糖轉化酶與活化因子的交互作用....33 第四章 結論與未來展望........................35 第一節 實驗結論..............................35 第二節 未來研究方向..........................35 2.1 探討活化因子與蔗糖轉化酶的交互作用 35 2.2 探討活化因子的生化性質 36 2.3 純化蔗糖轉化酶抑制因子 36 REFERENCE :....................................37 圖 與 表.....................................41 | |
dc.language.iso | zh-TW | |
dc.title | 綠竹 (Bambusa oldhamii) 蔗糖轉化酶活化因子之純化與檢定 | zh_TW |
dc.title | Purification and characterization of invertase activator from Bambusa oldhamii | en |
dc.type | Thesis | |
dc.date.schoolyear | 97-2 | |
dc.description.degree | 碩士 | |
dc.contributor.oralexamcommittee | 宋賢一(Hsien-Yi Sung),楊健志,林忠亮,張珍田 | |
dc.subject.keyword | 綠竹,蔗糖轉化酶,活化因子, | zh_TW |
dc.subject.keyword | Bambusa oldhamii,invertase,activator, | en |
dc.relation.page | 64 | |
dc.rights.note | 未授權 | |
dc.date.accepted | 2009-08-17 | |
dc.contributor.author-college | 生命科學院 | zh_TW |
dc.contributor.author-dept | 微生物與生化學研究所 | zh_TW |
顯示於系所單位: | 微生物學科所 |
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