(121h) Adsorption of Hydrophobically Modified DNA to Micelles, Liposomes, and Microemulsions

Methods to purify and concentrate DNA targets from complex mixtures are of great importance for identification of pathogens and forensic identification. A difficulty encountered when attempting to isolate DNA targets from these mixtures is that proteins, lipids, and other adventitious material tends to foul these surfaces, deactivating surface-attached oligonucleotide probes. We have been exploring the use of a tag-and-separate approach using DNA-binding surfactants that first bind targets in solution, then partition to micelles or other surfactant assemblies for separation. Here, we show that attachment of a single alkane to a DNA target up to 1000 bases in length allows for a significant but reversible adsorption to micelles, while no adsorption occurs for unmodified DNA.

In our approach, separations of tagged DNA from untagged DNA are performed by capillary electrophoresis. DNA samples are incubated with a peptide surfactant that hybridizes only to particular DNA sequences. A small aliquot of this solution is hydrodynamically injected into the capillary and then flushed with a solution of nonionic surfactant micelles (Triton X-100) by electro-osmosis. Tagged and untagged DNA are separated by virtue of the partitioning of the tagged DNA to the fast-moving micelles. Pseudo-equilibrium binding constants are obtained for the partitioning process by proper analysis of the elution times under various conditions.

We present a comprehensive description of the partitioning of hydrophobically tagged DNA to non-ionic micelles. We find that the partitioning process is a strong function of the DNA length, the length of the attached alkane, and the location of the alkane attachment along the DNA chain. We find a surprisingly strong dependence of the alkane chain length on the partitioning, with even a 2-carbon increment having a major impact. We also find that that type of micelle, liposome, or microemulsion used as an adsorbing phase drastically impacts the partitioning efficiency. Finally, we show that stretches of unbound DNA on either side of the target sequence do not interfere with the partitioning, making high-resolution separations of unprocessed DNA possible by this approach. We will also present preliminary data demonstrating high-resolution, sequence-specific DNA separations in the presence of large amounts of serum proteins.