Research of DNA stretching on lipid bilayer

本論文的目的在於開發具有良好的DNA分子拉伸效果，製程相對簡單且成本低的拉伸平台。現行眾多拉伸DNA分子的方法中，以侷限拉伸法的拉伸均一度為最佳，由於其原理為透過將DNA分子侷限於狹窄的空間(如：奈米通道、奈米狹縫等)中，並利用DNA分子自身不相交疊的特性來促使其延展並拉伸，因此DNA分子的延展度將隨著侷限空間的縮小而增加，縮小侷限空間便隨之成為發展趨勢；但同時卻也帶來了製程上的困難。為了能於拉伸效果不變的前提下，發展出更為優秀的拉伸平台，我們以侷限拉伸法的原理做為基礎，衍伸出嶄新的DNA拉伸方法：帶狀侷限法。 帶狀侷限法的核心概念是將DNA分子侷限於奈米尺度的帶狀平面，由於DNA為帶負電的高分子，我們將目標訂為製作出數百奈米寬的正電帶狀平面；假設DNA分子位於直角坐標系中，數百奈米的圖形寬度與靜電力的吸引將各自對DNA分子進行X軸與Z軸方向的侷限，促使DNA分子僅能延Y軸方向延展來達到被拉伸的目的。 正電帶狀平面的核心建立技術，共可分為兩個部分，一為如何建立均勻平整的帶正電平面，二為帶狀圖形的建立：使帶電平面達到我們數百奈米的寬度需求。首先，我們以架設於蓋玻片上的正電脂雙層(lipid bilayer)做為帶正電平面，帶狀圖形的建立則依方法原理的不同，衍伸出兩種較奈米通道簡易的製程：側壁限制法與帶狀圖形限制法。 利用側壁限制法，DNA分子的平均延展度可達到0.56，拉伸效果約相等於長寬介於60nm至80nm的矩形奈米通道，此結果證明了側壁限制法除了製程上的優勢，更具有優秀的拉伸效果。另一方面，以300nm寬的帶狀圖形限制法所獲得的DNA分子延展度約為0.4，其目前結果雖仍不及側壁限制法，但卻具有較側壁限制法更大的改善空間。帶狀侷限法目前於拉伸效果上雖仍不能取代現有侷限拉伸法，但相信能為相關領域帶來新的思維與方向。This thesis focuses on the development of a new platform that is capable of stretching DNA to a high degree while can be manufactured economically. Among current technologies for DNA stretching, confining DNA molecule in a nanochannel has been shown to provide the best stretching uniformity. “Confining DNA” means restricting DNA in a small space. Due to the excluded volume effects, DNA will extend more when the space becomes narrower. However, a reasonable DNA extension can only be achieved when the dimension of a nanochannel reduces to below 100nm, making such nanochannel expensive and difficult to produce. Inspired by the concept of confinement, we develop an alternative approach, called strip confinement, to stretch DNA molecule. The core concept of strip confinement is to restrict DNA in a strip-shaped two-dimensional plane. The two-dimensional nature of the platform enhances the excluded volume effects, and therefore is more advantageous for the application of DNA stretching. Since DNA is negatively charged in physiological condition, the proposed platform is realized by creating parallel strips of positively charged lipid bilayer on a glass. The processes for manufacturing our platforms, including the creation of the patterned surface and the setup of lipid bilayer, are more economical than those for producing nanochannels. Based on our new approach, we investigate two different ways to stretch DNA, sidewall confinement and direct strip confinement. The former stretches DNA along its corner where the lipid density is highest, the latter stretches DNA along its axis. The average degree of DNA extension measured from a sidewall confinement is 0.56, close to that obtained from nanochannels with the width between 60nm to 80nm. The degree of DNA extension measured from a Direct strip confinement with 300nm in width is 0.4, a little lower than that of the former method. The results presented here are for proof-of-principle, and we expect that our devices based on the concept of strip confinement can be further improved for better performance and lower cost