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研究生:陳惠萍
研究生(外文):Hui-Ping Chen
論文名稱:人類纖維母細胞株受低劑量輻射照射後基因表現之分析
論文名稱(外文):Microarray Gene Expression Analysis of Human Fibroblast Cell Line Exprosed to Low-Dose Radiation
指導教授:楊素珍楊素珍引用關係
指導教授(外文):S-J Yang
學位類別:碩士
校院名稱:中臺科技大學
系所名稱:放射科學研究所
學門:醫藥衛生學門
學類:醫學技術及檢驗學類
論文種類:學術論文
論文出版年:2007
畢業學年度:95
語文別:中文
論文頁數:73
中文關鍵詞:微陣列分析低劑量輻射基因異常
外文關鍵詞:low dose diagnostic X-ray irradiationgene expressioncDNA microarray analysis
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輻射線對於細胞的影響,較具規模的研究起始於1945 年原子彈爆發之後,一般認為輻射對人體造成影響,隨劑量增加嚴重度就越大。所以目前的研究資料,還是以累積高劑量的輻射生物效應資料比較齊全,觀察輻射對細胞產生的影響,最被熟知的方法,就是觀察染色體的變化,如斷裂或位移,或是較微觀的DNA 突變,近年來人類基因圖譜被解開,以微陣列分析方法分析細胞的基因表現也廣為被接受。回顧文獻我們發現,除了高劑量會對細胞造成影響外,低劑量輻射也會造成染色體不穩定及基因表現的改變,也有人曾利用微陣列分析法了解放射治療後病患細胞基因表現及生物標記特徵的變化。本實驗想了解低劑量輻射對細胞影響,並利用微陣列分析其中基因表現的變異,我們使用人類肺部纖維母細胞轉型(VA-13 )及
未轉型(MRC5)兩種細胞株,以醫用電腦斷層給予輻射照射0.1及0.25格雷之劑量,觀察其生長速率及外型的變化,並以微陣列分析實驗獲取可能因受低劑量輻射照射而可能會產生基因轉錄變異的基因。我們發現,不論轉型或正常細胞都因低劑量輻射照射而產生生長速率變慢的影響,並伴隨有大量基因表現改變的現象。本實驗分析15009 個的基因,微陣列分析結果,MRC5 細胞基因異常的比例為20.3%。VA-13 細胞基因異常的比例為6.5%。
In this study the effects of low dose diagnostic x-ray irradiation on human lung fibroblast cell lines were investigated. The cell growths of normal (MRC-5) and transformed (VA-13) lung fibroblasts were measured by MTT assay, and their associated genes expression profiles were monitored with cDNA microarray analysis. Compared with control experiments, MRC-5 showed decrease of cell growth up to 20% upon irradiation with 0.25 Gy of X-rxy , while VA-13 showed a more significant decrease to about 40%. In terms of the patterns of gene expression obtained from cDNA microarray analysis, MRC-5 cells had more differentially expressed genes (DEG) (3050) than VA-13 cells (968). The characteristic DEG’s in each cell type were further compared and their functions discussed.
目錄

表目錄…………………………………………………………………1
圖目錄…………………………………………………………………2
第一章前言……………………………………………………………3
第二章文獻回顧………………………………………………………8
2.1-1 單一低劑量輻射使基因不穩定的頻率增加…………………8
2.1-2 以微陣列為工具解開正常組織受輻射傷害之反應…………8
2.1-3 細胞經游離輻射照射後之轉譯反應…………………………9
第三章材料與方法……………………………………………………11
3.1 實驗材料與儀器…………………………………………………11
3.2 實驗方法…………………………………………………………15
3.2-1 細胞培養及繼代方法…………………………………………15
3.2-2 實驗流程………………………………………………………17
3.2-3 RNA 萃取……………………………………………………19
3.2-4 微陣列基因晶片分析………………………………………21
3.2-4.1 微陣列基因晶片雜合實驗…………………………………21
3.2-4.2 微陣列基因晶片資料讀取…………………………………23
3.2-5 MTT Assay……………………………………………………25
3.2-6 照射源設計及劑量計算………………………………………27
第四章結果…………………………………………………………30
4.1 MTT Assay 結果…………………………………………………30
4.2 微陣列基因晶片分析結果………………………………………33
4.2-1 MRC5 細胞直接照射基因變異之結果………………………33
4.2-2.1 MRC5 細胞直接照射後過度表現基因統計………………36
4.2-2.2 MRC5 細胞直接照射後表現過低基因統計………………38
4.2-3 VA-13 細胞直接照射基因變異之結果………………………40
4.2-4.1 VA-13 細胞直接照射後過度表現基因統計………………43
4.2-4.2 VA-13 細胞直接照射後表現過低基因統計………………45
4.2-5 直接輻射照射細胞異常基因統計……………………………47
4.2-6 直接輻射照射MRC5 及VA-13 細胞表現差異趨勢相同之異常基因比較…………………………………………………………………49
4.2-7 表現差異趨勢相反的異常基因比較…………………………58
第五章討論……………………………………………………………64
附錄……………………………………………………………………69
參考文獻………………………………………………………………70
參考文獻

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