Flowable networks as DNA sequencing media in capillary columns.

Abstract

A novel class of materials that self-assemble in water into equilibrium network structures with a well-defined mesh size consist of polyethylene glycols (PEG's) end-capped with micelle-forming fluorocarbon tails. These micellar systems form flowable aqueous gel-like networks that permit electrophoretic DNA sequencing in capillary columns. The gels have unusual rheological properties, including network breakdown under shear, resulting in plug flow that allows columns refill with complete ejection of byproducts of the previous sequencing analysis. In this system, DNA fragment electrophoretic mobilities are unaffected by the hydrophobicity of the polymer tails. Low molecular weight (M) PEG chains (M 8000) show catastrophic resolution loss for DNA fragments larger than 100 bases due to band broadening. For a longer PEG segment (M 35000) separating the end groups, band broadening occurs for DNA fragments larger than 300 bases, implying that the PEG segment length controls the mesh size in the equilibrium network structure. Optimum sequencing results were obtained from a 6% solution of a 1:1 mixture of C6F13 end-capped- and C8F17 end-capped PEG 35,000. The resolution limit of fluorescent-dye-labeled sequencing products in this formulation was 450 bases in 75 microns capillaries at 200 V/cm.