DNA gel electrophoresis is arguably the most widespread laboratory method in molecular biology, underlying an enormous range of analytical and preparative tasks. The speed and efficiency of this process could be greatly advanced by the DNA prism, a microfluidic method for continuously separating a mixture of DNA by size. This device provides (i) the molecular weight of the components (the analytical task) and (ii) purified components at the outlets (the preparative task). Existing DNA prisms were fabricated via photolithography and reactive ion etching of fused silica or by colloidal self-assembly. Unfortunately, neither of these methods is sufficient to move the DNA prism into routine use in biology labs. The long-term goal of this research program is to develop new nanostructured media that can provide order of magnitude improvements in separation speed and resolution when compared to gel electrophoresis. The objectives of this particular application are to (i) develop a process that is suitable for mass-production of DNA prism electrophoresis chips containing a large, three-dimensional array of perfectly ordered, uniform, sub-micron pores and (ii) demonstrate the ability of this medium to separate DNA. To accomplish these goals, an optical patterning method will be used to fabricate nanostructures in photoresist, the nanostructures will be integrated into a microchannel that provides a pulsed electric field, and the separating power of the device will be established using DNA ladders. To aid in the engineering of this device, optical theory will be used to design the structures and a Monte Carlo simulation model will be used to predict the separation from a particular pore structure. The research plan is thus divided into two specific aims:
Specific Aim 1 : Fabricate a DNA prism through optical patterning and demonstrate its separation ability.
Specific Aim 2 : Engineer optimal DNA prisms for separating different DNA size-ranges. This research is significant because it will lead to substantial reductions in the cost and time required for DNA separations. Optically patterned media should (i) exhibit marked improvements in separation time and resolution when compared to gel electrophoresis while (ii) being much easier to fabricate and more robust than existing nanofluidic devices for DNA separations. The devices produced by this research will impact molecular biology in general and genomics in particular by providing a route towards mass-produced chips for sizing DNA and collecting the products. The work is innovative because it uses methods developed in the field of photonic crystals to provide novel media for biomolecule electrophoresis. Taken as a whole, this research program will impact the larger field of biomicrofluidics and nanofluidics by providing approaches to create three-dimensional ordered media.
The proposed work will lead to improved nanoscale systems for rapid and high-resolution separations of DNA. These devices will accelerate a number of key genomics applications, such as DNA fingerprinting of infectious organisms and genome assembly.
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