One of the most time-consuming steps in DNA sequencing is the length-based, electrophoretic separation of enzymatically obtained copies of the sequencing template. Because DNA is a free-draining polymer, its electrophoretic mobility in free solution is not a function of length. As such, water-soluble polymers or gels are used as a sieving matrix, imposing obstacles upon the migrating DNA that retard its mobility in a length-dependent way. An alternative, "end-labeled free solution electrophoresis (ELFSE)," has been proposed in which a free-solution separation of DNA is realized by covalently attaching an uncharged polymer tag to the end of the DNA. The resulting end-modified DNA has a mobility intermediate between that of the polymer and unmodified DNA, whose value is a function of the DNA length. In principle, the method can provide separations that are 10-100 times faster than conventional methods. However, to be competitive with current techniques, the end-attached polymers must be extraordinarily monodisperse so that DNA oligomers differing by only a single base in length can be distinguished.

Here, we present a system in which surfactant micelles are transiently attached to DNA migrating in free-solution electrophoresis. DNA oligomers are alkylated prior to their enzymatic extension, then electrophoretically separated in buffers containing micelles of the nonionic surfactant Triton X-100. Micelle lifetimes are less than one second, and as such the alkylated DNA will exchange thousands of micelles during its migration. This thermal averaging effect confers a highly uniform drag upon all alkylated DNAs in the sample of interest as required for ELFSE. Furthermore, the self-assembling properties of micelles make them easy to incorporate into microfluidic devices and are compatible with substantial amounts of protein and lipid contaminants. Along with its application to rapid DNA sequencing, the micelle tagging method can be also be used to quantify mRNA levels and characterize surfactant microstructures.

We demonstrate that the transient attachment of alkylated DNA provides accurate base calling using commercial instruments. We also show that transiently attached micelles provide nearly the same drag as a covalently attached object having the same hydrodynamic radius. We will discuss our recent efforts to dramatically increase the lengths of DNA that can be analyzed in a short time using nanoemulsion droplets in place of micelles.