IMPROVED SEPARATION MEDIA FOR ELECTROPHORESIS

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In DNA sequencing analysis by means of electrophoresis, the separation medium could use further improvements. The long-term objective of the proposed project is to develop a better separation medium which can overcome many of the problems related to the present-day polyacrylamide solutions or gels: polymer solutions are often too viscous to be filled in capillary tubings; the polymerization in situ approach is not so reproducible; shelf-life for polymer gels is limited. The central theme is to take advantage of the self-assembly behavior of block copolymers so that predesigned polymer networks can be formulated by considering the chemical nature of the blocks, the total chain length, the block length ratio and the molecular architecture of the blocks in a selective solvent which is the buffer solution used in DNA sequencing analysis. In the self-assembled polymer networks, it is expected that several desirable properties can be designed into the properties of the separation medium. For examples, (1) at temperatures outside the operating temperature range for electrophoresis, the low molecular weight block copolymer (e.g. Pluronic polyols) solution has a low viscosity for ease of filling in capillary tubings or thin slabs, and (2) at the operating temperatures for electrophoresis, the self-assembled supramolecules can form gels of predetermined structures suitable for DNA sequencing analysis. In order to provide a basis for the design of a better separation medium, the structures of these entangled polymer chains (or gels) will be studied by means of physical techniques, such as viscosimetry, rheometry, laser light scattering, small angle neutron scattering, small angle x-ray scattering, C13 and proton NMR, and be correlated with electrophoresis results. By using the same physical techniques, it is proposed to study the effects of polydispersity on polymer chain entanglements and the use of a mixed polymer (e.g. polyacrylamide) solution/agarose gel approach which will be used to examine whether larger mesh size polymer networks can be achieved in order to resolve larger size DNA fragments. The agarose gel is used to restrict the translational motions of polyacrylamide chains so as to control the viscosity of the separation medium and the polymer chain dynamics.