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Please use this identifier to cite or link to this item: http://ntour.ntou.edu.tw:8080/ir/handle/987654321/10166

Title: 近場電紡微流體晶片製作奈米DNA纖維於原位雜交定序之應用
DNA Molecules Stretched and Embedded in Polymer Nanofibers by Near-Field Electrospinning Microfluidic Chip for Applications of Fiber- Fish Sequencing
Authors: 黃士豪
Contributors: NTOU:Department of Mechanical and Mechatronic Engineering
Keywords: DNA 纖維螢光原位雜交;近場電紡;奈米纖維
Fiber-FISH;Near-field electrospinning;nanofiber
Date: 2008-01
Issue Date: 2011-06-28T07:37:58Z
Publisher: 行政院國家科學委員會
Abstract: 摘要:DNA 纖維螢光原位雜交(Fiber-FISH, fiber based fluorescent in situ hybridization)技 術,是近年來發展起來的一項直接在DNA 纖維上進行螢光原位雜交的技術,其原理是 將具有螢光標記的DNA 探針直接原位雜交到DNA 纖維上,用來檢測DNA 序列在DNA 纖維上的空間定位。因此,本研究設計一近場電紡微流體晶片製作奈米DNA 纖維於原 位雜交定序之應用,藉由近場電紡技術(near-filed electrospinning process),將單一DNA 分子置入聚合物奈米纖維(nanofibers)內,產生直徑100 至350 nm 的奈米纖維。利用近 場電紡噴流中的展伸力可使DNA 分子幾乎完全伸展開來且彼此分離,再者,完全展伸 的DNA 分子嵌入在聚合物奈米纖維中,可永久維持展伸的構形且侷限在奈米尺度的聚 合物纖維空間內,可不受外界環境影響,再藉由電滲透流或是使用表面張力驅動流體方 式帶動具有DNA 分子嵌入的奈米纖維,以便觀察特定DNA 分子探針的序列。不同於 傳統電紡技術,採用固定或旋轉基板方式,本研究創新之處在於採用近場方式(探針和 收集板距離
abstract:Fiber-FISH is a technique recently developed to detect directly DNA sequences on extended DNA fibers by fluorescence in situ hybridization (FISH), the resolution and sensitivity of which are improved to 1-2 kb and 200 bp respectively. A rapid molecular mapping technology—Direct Linear Analysis (DLA)—on the basis of the analysis of individual DNA molecules bound with sequence-specific fluorescent tags is a practicable method. In this study, we propose a near-field electrospinning microfluidic chip for applications of Fiber- Fish sequencing. Based on the near-field electrospinning technique, the proposed microfluidic chip can encapsulate single molecules of DNA inside polymer nanofibres by an electrospinning process. The DNA molecules were stretched to nearly their full length. The resulting nanofibers were 100 - 350 nm in diameter. The strong elongational forces present in electrospinning jets could serve to stretch DNA into their full length. The stretched DNA molecules encapsulated in a protective nanofiber can be subsequently collected, manipulated, and driven by electroosmotic flow or surface-tension flow to optically analyze the probe sequences. Instead of the conventional electrospinning technique using a conductively rotational plate for collection, we use the near-field electrospinning technique, i.e. the distance between collected plate and probe below 1 cm, and microfluidics to collect and transport nanofibers to the optical detection area at 10μm/s. A detection system capable of single-fluorophore sensitivity is used to detect at sequence-specific motif sites on the stretched DNA molecules. DLA provided the spatial locations of multiple specific sequence motifs along individual DNA molecules, and thousands of individual molecules could be analyzed per minute. Besides, by using the techniques of surface-patterned modification and microfluidic self-alignment, we can align nanofibers into a functional structure for the subsequently optical detection. The proposed near-field electrospinning microfluidic chip for applications of Fiber- Fish sequencing is a quick, inexpensive, and straightforward method to obtain many stretched molecules allowing for single molecule analysis, which can be a useful tool in human genome mapping.
Relation: NSC97-2218-E019-001-MY2
URI: http://ntour.ntou.edu.tw/ir/handle/987654321/10166
Appears in Collections:[機械與機電工程學系] 研究計畫

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