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研究生: 謝磊
Hsieh, Lei
論文名稱: 利用iTRAQ化學標定方法進行乳癌細胞系MCF-7多重抗藥性之差異蛋白質體學分析
Differential proteomic analysis of multidrug resistance in breast cancer cell line MCF-7 by iTRAQ technology
指導教授: 陳頌方
Chen, Sung-Fang
學位類別: 碩士
Master
系所名稱: 化學系
Department of Chemistry
論文出版年: 2020
畢業學年度: 108
語文別: 中文
論文頁數: 99
中文關鍵詞: 差異蛋白質乳癌
英文關鍵詞: differential protein
DOI URL: http://doi.org/10.6345/NTNU202001590
論文種類: 學術論文
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  • 謝誌 i 中文摘要 ii Abstract iii 目錄 iv 表目錄 vii 圖目錄 viii 第一章 緒論 1 第一節 乳癌 1 第二節 乳腺癌細胞系MCF-7 1 第三節 乳癌荷爾蒙療法介紹 2 第四節 乳癌治療藥品 3 一、 Tamoxifen (泰莫西芬) 3 二、 Everolimus 4 三、 Palbociclib 6 第五節 質譜儀技術 7  高能碰撞解離(High energy collision dissociation, HCD) 8  軌道阱(Orbitrap)質量分析器 9 第六節 蛋白質身份鑑定 11 第七節 差異蛋白質體學 13 第八節 液相層析分離技術 22 一、 鹼性逆相層析法(Basic reverse phase chromatography, bRP) 23 二、 強陽離子交換層析法(Strong cationic exchange chromatography) 24 三、 液相等電點聚焦分離法(Solution isoelectric focusing) 25 第九節 研究動機 27 第二章 實驗材料與方法 29 第一節 樣品介紹 29 第二節 樣品製備 29 一、 四株抗藥性細胞株蛋白收集 30 二、 收集 medium 蛋白 31 第三節 樣品純化濃縮 31 第四節 蛋白質濃度測定 (Bradford protein assay) 32 第五節 蛋白質水解(In-solution digestion)和化學標定iTRAQ®試劑 33 第六節 第一維分離策略 35 一、 鹼性逆相層析法(Basic reverse phase chromatography, bRP) 35 二、 強陽離子交換層析法(Strong cationic exchange chromatography) 36 三、 液相等電點聚焦分離法(Solution isoelectric focusing) 38 第七節 自製碳18離心管柱 (C18 spin column) 去鹽 40 第八節 奈米級超高效能液相層析電噴灑串聯式質譜儀 (nanoLC ESI tandom mass spectrometry) 42 一、 超高效能液相層析 (Ultimate system 3000 nanoLC system, Thermo Fisher Scientific ) 42 二、 Orbitrap Fusion Lumos 軌道阱質譜儀 43 第九節 資料分析 (Data analysis) 46 第三章 結果與討論 48 第一節 蛋白質樣品濃度測定 48 第二節 iTRAQ 試劑標記 51 第三節 蛋白質身份鑑定及相對定量 52 第四節 鹼性逆向層析法分離樣品之質譜鑑定結果 53 第五節 強陽離子交換層析法分離樣品之質譜鑑定結果 57 第六節 等電聚焦分級分離儀分離樣品之質譜鑑定結果 61 第七節 比較三種不同分離方法之蛋白質及胜肽鑑定結果 64 第八節 不同樣品蛋白質相對定量 72 第九節 蛋白質相關生物作用途徑 77 第十節 相關差異蛋白質探討 84 第四章 結論與未來展望 89 第五章 文獻參考 90

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