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研究生:李俊彥
研究生(外文):JUAN-YAN Li
論文名稱:矽晶片上成長之聚異丙基丙烯醯胺高分子刷藉由熱感應轉換特性作為萃取人體DNA
論文名稱(外文):Extraction of human DNA by poly(N-isopropyl acryl amide) brushes on the silicon surface through thermally responsive switching
指導教授:陳建光陳建光引用關係
指導教授(外文):Jem-Kun Chen
學位類別:碩士
校院名稱:國立臺灣科技大學
系所名稱:高分子系
學門:工程學門
學類:化學工程學類
論文種類:學術論文
論文出版年:2009
畢業學年度:97
語文別:中文
論文頁數:97
中文關鍵詞:原子轉移自由基聚合反應DNA萃取聚合酵素鏈鎖反應聚異丙基丙烯醯胺高分子刷
外文關鍵詞:poly(N-isopropyl acryl amide) brushesATRPDNA exactractionPolymerase chain reaction
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藉由原子轉移自由基聚合反應將聚異丙基丙烯醯胺(poly N-isopropyl acryl amide, PNIPAAM)高分子刷接枝於矽晶片表面。晶片表面經過氧電漿處理後可增加表面氫氧基的含量,而表面的氫氧基可與3-aminopropyltriethoxy silane(APTES)反應,接著用α-Bromoisobutyl bromide與接枝上表面的APTES反應,以作為表面接枝高分子的起始端,藉由原子轉移自由基聚合反應(atom transfer radical polymerization, ATRP)可合成PNIPAAM高分子刷於矽晶片表面。X光光電子能譜(X-Ray photoelectron Spectroscopy,XPS)用於分析表面自組裝起始端以及PNIPAAM高分子之表面元素,使用橢圓測厚儀(Ellipisometer)量測PNIPAAM高分子刷厚度,而 PNIPAAM高分子刷在經過2小時的高分子聚合其厚度達到71nm,並使用原子力顯微鏡AFM(atomic force microscope)分析表面狀態。AFM影像顯示出在30-120分鐘聚合時間下之表面形態,在120分鐘的接枝時間均方根粗糙度為5.2537nm。

由由接觸角的量測,由接觸角60o變化80o,我們可以明顯的觀察到調控的親水疏水性質變化。我們證實了在反覆的低臨界濃度溶解溫度(Lower Critical Solution Temperature, LCST)循環測試下,證明其PNIPAAM高分子刷之可靠度。

利用PNIPAAM高分子刷之親疏水轉換機制,藉由聚合酵素連鎖反應(Polymerase Chain Reaction ,PCR)以及凝膠電泳(Gel electrophoresis研究人體血液染色體的去氧核醣核酸(Human blood genomic DNA)抓取釋放實驗,


實驗觀察指出,PNIPAAM高分子刷會在LCST溫度以下抓取50ng/ul 以及133ng/ul的特定DNA分子,在LCST溫度以上釋放出DNA分子。由結果得知,可以利用在矽晶片基材上面成長的PNIPAAM高分子刷作為一種新的特定DNA的萃取方式。
The poly (N-isopropylacrylamide) (PNIPAAm) brushes were grafted on the silicon surface by using atom transfer radical polymerization (ATRP). The silicon wafer surface was treated by oxygen plasma to increase the hydroxyl groups. The hydroxyl group on the surface reacted with 3-aminopropyltriethoxy silane (APTES), then α-Bromoisobutyl bromide was used to reacted with APTES to form the initiator of grafting polymerization on the surface. The PNIPAAm brushes was then synthesized on the surface through ATRP. The X-Ray photoelectron Spectroscopy(XPS) was utilized to analyze the surface element of SAMs of initiator and PNIPAAm brushes. The thickness of the PNIPAAm brushes on the surface was about 71 nm for 2 hours of polymerization time from mensurement by ellipisometer. The morphology of PNIPAAM was investigated by atomic forece microscope (AFM). AFM images show the morphology of the PNIPAAM brushes in 30-120 minutes polymerization, and The Root Mean Square roughness of PNIPAAM brushes in 120 polymerization time was 5.2537nm.

The surface with PNIPAAm brushes demonstrated obvious variations of hydrophobic and hydrophilic properties in contact angle measurement from 60o to 80o. We verify the reliability of PNIPAAm brushes through the 3 of the LCST cycle tests.

According to the hydrophobic and hydrophilic properties of PNIPAAM brushes, the capture-release application for human blood genomic DNA was investigated by Polymerase Chain Reaction(PCR) and Gel electrophoresis.

The observation suggests that the PNIPAAM brushes captured the human specific DNA form 50ng/ul and 133ng/ul of solution as the temperature below LCST. And the DNA was released as the temperature above LCST. The results demonstrated the new strategy for extraction for specific DNA by PNIPAAM brushes on the silicon substrate.
摘 要 I
Abstract III
致謝 V
表目錄 X
圖目錄 XI
第1章 前言 1
1.1 研究背景 1
1.2 研究目的 2
第2章 文獻回顧 4
2.1 高分子刷發展與應用回顧 4
第3章 相關實驗理論 17
3.1高分子材料製成 17
3.1.1原子轉移自由基聚合法 17
3.1.2 表面起始聚合 21
3.1.3 感熱型高分子-聚異丙基丙烯醯胺 23
3.2 材料表面檢測分析 24
3.2.1原子力顯微鏡 24
3.2.2 X光光電子能譜 30
3.2.3接觸角量測 33
3.3生物技術與量測原理 35
3.3.1聚合酵素鏈鎖反應 35
3.3.2 凝膠電泳Gel electrophoresis 40
第4章 實驗設備與方法 42
4.1實驗材料 42
4-2實驗儀器 43
4-3 矽晶片上高分子刷接枝製備 45
4.4 生物實驗測試 48
4.4.1 聚合酵素連鎖反應試驗 48
4.4.2 生物分子抓取釋放實驗 53
第5章 結果與討論 57
5.1 矽晶片上高分子刷之製備結果分析 57
5.1.1 XPS, X-ray Photoelectron Spectroscopy分析 57
5.1.1.1 N 1s圖譜分析 58
5.1.1.2 Br 3d圖譜分析 59
5.1.1.3 C2p圖譜分析 59
5.1.1.4 Si2p圖譜分析 60
5.1.2 高分子刷之厚度分析 62
5.1.3 接觸角量測分析 63
5.1.4 AFM表面型態分析 66
5.2 生物分子分析 68
5.2.1 PNIPAAM高分子刷抓取試驗XPS分析 68
5.2.2 生物PCR成品電泳照膠分析 69
5.2.2.1 Human blood g-DNA針對引子PCR電泳照膠分析 69
5.2.2.2抓取釋放Human blood g-DNA PCR電泳照膠分析 72
第6章 結論 74
參考文獻 75
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