利用電場調控聚電解質多層膜的釋放 以應用於基因輸送

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姓名

鍾坤達(Kun-da Chung) 查詢紙本館藏

畢業系所

化學工程與材料工程學系

論文名稱

利用電場調控聚電解質多層膜的釋放以應用於基因輸送
(The manipulation of gene delivery from PEI/DNA multilayers by the application of electrochemical potentials)

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摘要(中)

摘要
在本次實驗中，利用聚乙烯亞胺(Polyethylenimine)和DNA在具有生物相容性的導電基材聚吡咯(Polypyrrole) 上面進行疊層組裝，並將所形成多層膜之結構進行通電釋放。從釋放實驗結果顯示以固定縱向通電的方式可以有效的從膜中釋放出PEI和DNA，在進一步的探討中，電壓成為了主要影響釋放的參數，通予越大電壓，釋放出的PEI和DNA的比例就越高。對於增加通電時間，也有類似的效果。進一步將通電過後的多層膜，以原子力顯微鏡去拍攝，比較有通電與沒通電之結果，可以發現，通電過後膜的表面粗糙度有了明顯的上升，通電越大，粗糙度相對就越大，可能原因是由於膜的厚度不均，因電場和電化學作用下，導致膜比較薄區域，越容易溶解，導致膜高低起伏會隨之增加。最後將所釋放的DNA及PEI進行轉染實驗，可發現通電組其轉染效率高於無通電組，證實所釋放的DNA及PEI，可以有效地被細胞所攝取並提升了轉染效率。

摘要(英)

Abstract
In this study, polyethylenimine (PEI) and DNA were deposited onto conductive polypyrrole (PPy) substrate using layer-by-layer assembly. The formed polyelectrolyte multilayers (PEMs) films were released under electric field. The release results suggested that the electric field perpendicular to the substrate can efficiently release PEI and DNA from PEMs. Voltages were the main factors controlled the PEMs disassembly that higher potential released more PEI and DNA. The treatment period demonstrated similar effects. The surface morphologies of PEMs after electric field treatment were observed using atomic force microscopy (AFM). Compared to the nontreated group, the roughness of the films significantly increased after treating electric field, which was more obvious as the potential was high. These results suggested that the polyelectrolytes may be easily released from thin regions of the films due to the driving force of electric field as well as the dissolution resulted from electrochemical reaction on electrodes. Finally, the release DNA and PEI were used for transfection. The released material from the films with electric field treatment demonstrated superior transgene expression, suggesting that the electric assisted gene delivery from PEMs should be beneficial to gene delivery.

關鍵字(中)

★ 疊層組裝
★ 電場
★ 釋放
★ 聚吡咯
★ 基因
★ 聚乙烯亞胺

關鍵字(英)

★ Layer-by-Layer
★ Electric field
★ release
★ Polypyrrole
★ DNA
★ Polyethylenimine

論文目次

目錄
摘要 I
Abstract II
目錄 III
圖目錄 VI
第一章序論 1
1-1背景 1
1-2實驗目的 3
第二章文獻回顧 4
2-1組織工程 4
2-2基因治療 5
2-2-1基因治療的載體 5
2-3疊層組裝 8
2-3-1疊層組裝的原理 8
2-3-2疊層組裝的製備 8
2-3-3疊層組裝的在生物上的應用 9
2-3-4通電對疊層組裝釋放的影響 11
2-4聚吡咯(Polypyrrole) 21
2-5聚乙烯亞胺(Polyethylenimine) 22
2-5-1聚乙烯亞胺於基因傳遞之應用 22
2-6紫外光光譜儀 24
2-6-1紫外光光譜儀 24
2-6-2紫外光光譜儀原理 24
2-7原子力顯微鏡 27
2-7-1原子力顯微鏡原理 27
2-7-2原子力顯微鏡操作模式 28
第三章實驗方法 29
3-1實驗架構 29
3-2實驗藥品與儀器 30
3-2-1藥品 30
3-2-2儀器 30
3-3試藥製備 32
3-3-1 DNA cloning 32
3-3-2陽離子水溶液 33
3-3-3陰離子水溶液 33
3-3-4 Polypyrrole 製備 34
3-3-5 TNBSA溶液 34
3-4疊層組裝製備(LBL) 34
3-4-1基材製備 34
3-4-2疊層組裝 35
3-5 釋放實驗 36
3-5-1 DNA釋放量檢測 36
3-5-2 PEI釋放量檢測 36
3-6 HEK293T細胞轉染實驗 37
3-7 原子力顯微鏡分析 37
3-8 掃描式電子顯微鏡分析 37
第四章結果與討論 38
4-1 通電對聚電解質釋放影響 38
4-1-1 不同電場方向對聚電解質釋放影響 38
4-1-2 不同電流對聚電解質釋放影響 41
4-1-3 不同電壓對聚電解質釋放影響 43
4-1-4 通電不同時間對聚電解質釋放影響 46
4-2原子力顯微鏡探和掃描式電子顯微鏡探討表面結構變化 49
4-3細胞轉染 53
第五章結論 55
第六章參考文獻 56

參考文獻

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指導教授

胡威文

審核日期

2014-8-20

推文