以電腦模擬改良圓柱陣列微流道應用於DNA分離之研究

Abstract

在具有圓柱陣列的微流道中，若施加電場或流場，溶液中的DNA就被驅動並機率性地與微流道中的圓柱障礙物進行碰撞、上鉤與脫鉤。若利用分子量不同之DNA與圓柱障礙物的碰撞機率與脫鉤時間不相同的特性，便能達到分離之效果。本研究使用電腦模擬，利用布朗動態法(Brownian Dynamics) 連結有限元素法(Finite Element Method)以模擬DNA在微流道中運動之行為，探討透過圓柱陣列分離不同分子量DNA之效果。本研究中提出四大類具有圓柱陣列之微流道設計，用來分離λ-DNA (48.5 kbp) 與T4-DNA (165.6 kbp)，每一個模型皆是根據前者發現之問題進行改良，希望找出最佳之分離裝置。 一般來說，在高電場下DNA會受到通道效應(Channeling Effect)影響，大幅降低DNA與圓柱之碰撞機率，導致DNA無法分離。因此我們首先在圓柱陣列中加入漸擴通道，希望藉由在y方向的電場梯度產生預拉伸DNA之效果以增加碰撞機率，並由初步的模擬結果發現此方法確實能夠分離DNA。但引入漸擴通道的設計後， DNA之電泳之路徑較為分散而導致DNA通過每個分離單元之時間大不相同，增加了DNA分布之標準差，反而降低了分離解析度。因此我們改用流場做為驅動力，藉由流體動力作用所產生之空乏層(depletion layer)以限制DNA行經的路徑，有效改善前述問題。接著我們將通道狹窄區之長度縮短，以便在相同長度下置入更多分離單元，進一步增加裝置之分離效率。然而在實驗上，DNA被發現容易卡在通道狹窄區，且DNA之起始分布太廣，導致此設計在實作上無法使用。為了克服上述之問題，我們將漸擴與漸縮通道並聯，藉此增加通道狹窄區之寬度。我們發現本模型中電場會在漸擴區施予DNA一靠牆之合力，使其貼近牆壁電泳，能降低DNA分布的標準差，對高分子量的T4-DNA尤其顯著。此外，我們也分析在不同Pe下脫鉤時間與運動時間對DNA分布之標準差的影響，發現在低Pe時為電泳路徑主導，高Pe則轉變為脫鉤時間所影響。而此模型成功排除在高電場下之通道效應，保有良好的分離解析度，達到在短時間內分離DNA之效果，且隨著通道長度增加，整體裝置的解析度約與通道長度的0.5次方成正比。與實驗相比，雖然仍有相當誤差，但可以相當準確地預測其變化趨勢，並提供一個合理的實驗設計，大幅減少實驗開發的成本。

Microchannel with hexagonal post arrays is one of the novel devices designed for rapid separation of very long DNA by electrophoresis. However, its efficiency is poor at high Peclet number (or electric field) due to the channeling effect which reduces the probability of collision between DNA and posts. To overcome this drawback, we propose four different design of microchannels and test their ability for separating two model DNA, namely λ-DNA (48.5 kbp) and T4-DNA (165.6 kbp), by using Brownian Dynamics simulation in conjunction with finite element method. In the first design, we add a microexpansion in front of the post arrays so that the collision probability between DNA and posts increases due to the pre-stretching of DNA at the microexpansion. However, the separation power was found only increasing marginally due to the increasing variation of the available path for DNA electrophoresis. In the second and the third design, we replace electric field with flow field to reduce the variety of DNA path by a depletion layer caused by hydrodynamic interaction. However, in experiments DNA were found stuck easily at the narrow part of the channel. Moreover, the broad DNA injection band leads to a significant drop in separation resolution. In the last design, we connect the first design in parallel in order to reduce both the width of the injection band and the chance of channel clog. We also found DNA move toward convexed wall due to a normal force, and this phenomenon results in a reduction in DNA path. With this design, we obtain a good separation resolution in short time based on our simulation. We also investigated the factors that contribute to the standard deviation of DNA distribution. We found the variation of DNA path dominates at low Pe but the DNA unhooking time dominates at high Pe. Thus, it is important to control both factors for further improvement of the device. To conclude, we have used simulation to help improving the design of microchannel with post arrays for the purpose of DNA separation.