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Student Number 91324015
Author Li-An Kao(高立安)
Author's Email Address No Public.
Statistics This thesis had been viewed 2352 times. Download 2379 times.
Department Chemical and Materials Engineering
Year 2003
Semester 2
Degree Master
Type of Document Master's Thesis
Language zh-TW.Big5 Chinese
Title Studies of the Effect of Secondary Structure of single-stranded DNA on the Kinetics and Mechanism of Hybridization
by Surface Plasmon Resonance
Date of Defense 2004-06-04
Page Count 114
Keyword
  • oligo-nucleotide hybridization
  • secondary structure
  • surface plasmon resonance
  • Abstract This study utilizes a surface plasmon resonance (SPR) biosensor and a theoretical secondary structure calculation program(Oligo) to investigate the influence of secondary structures of ssDNA on the DNA hybridization. It is found that the SPR angular shifts associated with the
    three pairs of 60mer oligo-nucleotides with prominent secondary structures are lower than those observed for the two pairs of oligonucleotides with no obvious secondary structures. It is also determined that increasing the DNA hybridization temperature from 35 oC to 45 oC reduces secondary structure effects on hybridization. On the hybridization with mixture target oligo-nucleotides, the SPR results demonstrate that presence of oligo-nucleotides mixture due to non-specific interactions
    between non-complementary probes and targets interfere the extent of kinetics of hybridization significantly.
    In this investigation, thermodynamics information was obtained by Van’t Hoff equation and combining with the kinetics data from SPR, the
    hybridization mechanism were proposed and discussed.
    Table of Content 第一章前言...................................................................1
    第二章文獻回顧...............................................................3
    2.1 表面電漿共振(Surface Plasmon Resonance,SPR) ....................3
    2.1.1 表面電漿現象原理.........................................................3
    2.1.2 光學激發表面電漿之方式..................................................4
    2.1.3 SPR 檢測生物反應.........................................................7
    2.1.4 表面電漿共振光譜儀於分子交互作用上可提供之資訊.................8
    2.1.5 其他類型之表面電漿共振..................................................9
    2.2 SPR 於生物檢測上之應用領域................................................15
    2.2.1 蛋白質變(復)性研究......................................................15
    2.2.2 DNA 與Protein 之間的交互作用.....................................16
    2.2.3 Epitop Mapping..........................................................18
    2.2.4 抗原(Antigen)與抗體(Antibody)與之交互作用..............19
    2.2.5 生物膜(membrane)與酯雙層(lipid bilayer)......................20
    2.2.6 阿茲海默症致病研究(Alzheimer) ....................................22
    2.2.7 高分子膜(polymer films)研究..........................................23
    2.2.8 非特異吸附生物分子交互作用-生物相容性(biocompatibility)...............24
    2.2.9 組織工程...............................................................25
    2.3 去氧核醣核酸及核酸雜交反應................................................28
    2.3.1 去氧核醣核酸分子簡介....................................................28
    2.3.2 核酸雜交反應...........................................................30
    2.3.3 單股核酸之二級結構.....................................................36
    第三章實驗藥品與儀器設備.....................................................38
    3.1 實驗藥品.................................................................38
    3.2 實驗儀器設備..............................................................40
    3.2.1 表面電漿共振儀(Surface Plasmon Resonance) ...............40
    3.2.2 Oligo 序列模擬軟體.....................................................44
    3.3 實驗目的..............................................................46
    3.4 實驗方法..................................................................46
    3.4.1 實驗溶液配製..........................................................46
    3.4.2 實驗步驟............................................................47
    第四章結果與討論..........................................................51
    4.1 Oligo 模擬核酸序列之結果..............................................51
    4.2 二級結構對核酸雜交的影響.............................................59
    4.2.1 具明顯二級結構雜交實驗之結果....................................59
    4.2.2 非明顯二級結構雜交實驗之結果....................................61
    4.3 溫度效應對二級結構核酸雜交之影響.....................................62
    4.3.1 溫度效應對具明顯二級結構於核酸雜交反應之影響...62
    4.3.2 溫度效應對非明顯二級結構於核酸雜交反應之影響...65
    4.4 雜交反應動力學分析........................................................67
    4.4.1 反應模式的選擇........................................................67
    4.4.2 動力曲線擬合............................................................70
    4.4.3 擬合動力學參數之合理評估............................................75
    4.4.4 動力學分析...........................................................77
    4.5 混合樣品之雜交反應.......................................................80
    4.5.1 實驗設計................................................................80
    4.5.2 混合樣品之雜交反應................................................80
    4.6 雜交反應熱力學參數分析..................................................85
    第五章結論...............................................................87
    Reference ..................................................................90
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    Advisor
  • Wen-Yih Chen(陳文逸)
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    Date of Submission 2004-06-17

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