DNA在微通道的熱泳行為｜國立中央大學博碩士論文系統

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研究生：	林世彬 Shin-pin Lin
論文名稱：	DNA在微通道的熱泳行為 Thermophoresis of DNA in micro-channel
指導教授：	陳志強 Chan-chi Keung 黎璧賢 Pik-yin Lai
口試委員:
學位類別：	碩士 Master
系所名稱：	理學院 - 物理學系 Department of Physics
畢業學年度：	98
語文別：	英文
論文頁數：	98
中文關鍵詞：	熱泳、核糖核酸
外文關鍵詞：	Thermophoresis, Soret coefficient, DNA
相關次數：	點閱：16 下載：0
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我們研究400bp DNA, 48kbp λ-DNA 和166kbp T4 DNA在微通道中的熱泳運動。我們使用近紅外線雷射當作在微通道產生溫度梯度的加熱源。利用YoYoY-1螢光染劑和DNA結合後會產生1000倍強度的螢光特性，以螢光強度變化代替DNA的濃度變化並記錄下其在不同時間的影像。實驗後由所記錄的DNA影像進而分析出其量變曲線。最後再由量變曲線隨時間的變化以數值分析的方法量測得擴散常數和ST*。我們定義ST*=ΔT0ST。ΔT0是加熱點在加熱後和微通道兩端的溫度差。ST是Soret係數，定義為熱擴散系數與擴散系數之比。
這篇論文的目標旨在研究不同管徑的毛細玻璃管、不同長度的DNA、不同強度的加熱源、不同濃度的奈米等級的金粒子和不同濃度的亞精胺對DNA在熱泳實驗中的影響。我們在實驗中發現a)擴散常數和ST*的大小會和毛細玻璃管管徑和DNA回轉半徑的比例有關。b)當毛細玻璃管管徑比DNA回轉半徑大時，加熱源的強度並不會影響擴散常數和ST的大小。c)亞精胺會增加DNA的擴散常數但ST的大小卻會因為亞精胺的增加而減少。d) 奈米等級的金粒子會增加DNA的擴散常數但是對於ST似乎並無影響。

The motion of DNA in micro-channels due to thermophoresis is studied. 48kbp λ-DNA, 166kbp T4-DNA and 400bp DNA are used. They are tagged by fluorescent YoYo-1 molecules and placed in a micro-sized channel. An infrared laser is used to heat up the system forming a temperature gradient inside the channels. The DNA concentration gradient is then measured by recording the fluorescence intensity profiles of the YoYo-1. The measured DNA concentration profiles are found to be in good agreement with our numerical model fitting in which diffusion and thermophoretic fluxes are considered. Both the apparent Soret coefficient (ST*) and diffusion constant of DNA can be obtained from these numerical model fittings. We define the ST*=ΔT0ST.ΔT0 is the temperature rise of the heating spot. ST is Soret coefficient defined as ST=DT/D when DT is the thermal diffusion coefficient and D is the usual self-diffusion constant.
The purpose of this thesis is to study the effects of different capillary tube diameters, lengths of DNA, heating powers, concentration of nano-gold and concentration of spermidine on the properties of DNA thermophoresis. We find that a) diffusion constant and apparent Soret coefficient depend on the ratio of diameter of capillary tube to radius of gyration of DNA b) diffusion constant and Soret coefficient are independent of heating powers when diameter of capillary tube is bigger than radius of gyration of DNA. c) Spermidine will increase the diffusion constant of the DNA and decrease Soret coefficient with the increase of the concentration of spermidine d) Nano-gold will increase the diffusion constant of the DNA but have little effect on the apparent Soret coefficient with the increase of the concentration of nano-gold.

Chapter 1 Introduction………………………………………………………………1
1 Thermophoresis…………………………………………………………………1
1.1 Overview…………………….……………………………………………1
1.2 The physical property of thermophoresis……………………………..2
2 Thermophoresis of polymer…………………………………………………....4
2.1 Characteristics of polymer………………………………………………4
2.2 Physical properties of polymers………………………………………..5
2.3The phenomenon of polymer thermophoresis………………………...5
3 Thermophoresis of DNA………………………………………………………...6
3.1 The structure of DNA…………………………………………………….6
3.2 The phenomenon of DNA thermophoresis…………………………….7
3.3 The effect of temperature for the DNA thermophoresis………………9
4 Summary………………….……………………………………………………...10
Chapter 2 Setup and procedure…………………………………………………….12
1 Overview………………………………………………………………………….12
2 Introduction to fluorescent material…………………………………………….12
2.1 Fluorescent material: Oxazole Yellow dimmer (YOYO-1) …………...12
3 DNA sample preparation………………………………………………………..14
3.1 Introduction to materials…………………………………………………14
3.2 Photo-bleaching and protection solution………………………………14
3.3 Preparation for DNA-YOYO-1 complexes…………………………….15
3.4 Introduction to capillary glass tube…………………………………….15
4 Microscope……………………………………………………………………….16
4.1 Introduction to microscope……………………………………………...16
4.2 Principle of fluorescence microscopy………………………………….16
5 Continuous wave infrared laser………………………………………………..18
5.1 Introduction to continuous wave infrared laser………………………...18
5.2 The working principle of optical pumping……………………………....19
5.3 Infrared heating…………………………………………………………....19
6 Intensified CCD camera……………………………………………………….…20
6.1 Intensified CCD camera: PTI IC-200…………………………………....20
7 Optical setup (optical tweezers) ………………………………………………..22
7.1 An introduction………………………………………………………….….22
7.2 The setup for the laser tweezers…………………………………….…..22
8 Numerical model fitting for thermophoresis……………………………….……24
8.1. The model system…………………………………………………..……..24
8.2. The steps and methods of dynamics fitting………………………….…26
Chapter 3 Results and Discussion…………………………………………….……..30
1 Methods of Experimental data analysis………………………………….……..30
1.1 The raw picture and profile………………………………………….……30
1.2 The numerical model fitting………………………………………….……32
2 Results for different tubes diameters……………………………………………33
2.1 The raw pictures and profile of DNA ……………………………………33
2.2 Characteristics of the profile ………………………………………….….37
2.3 Time dependence of the depletion minimum……………………….…..37
2.4 The profile of the DNA for different heating powers……………………38
2.5 The diffusion constant……………………………………………….…….39
3 Results for different heating power……………………………………….……..39
3.1 The raw pictures and profile of DNA………………………….….………40
3.2 Characteristics of the profile ………………………………….….……....43
3.3 Time dependence of the depletion minimum……………….…….…….43
3.4 The profile of the DNA for different diameters……….………….……...44
3.5 Diffusion constant………………………………………………………..44
4 Numerical model fitting………………………………………………………….46
4.1 The dynamic numerical fitting model…………………………………..46
4.2 The results of model fitting for different capillary tube diameters…...60
4.3 The results of model fitting for different heating powers……………..63
5 Application of the method to some interesting cases…………………………..66
5.1 The comparison of λ-DNA and 400bp DNA in different capillary
glass tubes………………………………………………………………..66
5.2 The comparison of λ-DNA and T4 DNA in different heating powers..68
5.3 Effect of spermidine………………………………………………………70
5.4 Effect of nano-gold………………………………………………………..73
Chapter 4 Summary…………………………………………………………………..77
Reference………………………………………………………………………………79
Appendix……………………………………………………………………………….83
A Sample preparation for protection solution…………………………………83
B Sample preparation for DNA-YOYO-1 complexes solution………………84
C Laser-blocking device………………………………………………………..85
D program………………………………………………………………………..87

                                

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