Compression and self-entanglement of single DNA molecules under uniform electric field

Abstract

We experimentally study the effects of a uniform electric field on the conformation of single DNA molecules. We demonstrate that a moderate electric field (∼200 V∕cm) strongly compresses isolated DNA polymer coils into isotropic globules. Insight into the nature of these compressed states is gained by following the expansion of the molecules back to equilibrium after halting the electric field. We observe two distinct types of expansion modes: a continuous molecular expansion analogous to a compressed spring expanding, and a much slower expansion characterized by two long-lived metastable states. Fluorescence microscopy and stretching experiments reveal that the metastable states are the result of intramolecular self-entanglements induced by the electric field. These results have broad importance in DNA separations and single molecule genomics, polymer rheology, and DNA-based nanofabrication.DNA conformation | electrophoresis | polymer physics | knots | microfluidic T he use of electric fields to transport DNA has become an essential technique for a variety of research areas including molecular biology, gene therapy, and fundamental studies of polyelectrolytes (1). In most applications, nonlinear electrokinetics (2) due to the interplay of negatively charged DNA and the electric field are neglected. However, Viovy and coworkers (3, 4) demonstrated the ability of an electric field on order of 100 V∕cm to induce strong intermolecular DNA aggregation in a standard electrophoresis buffer in the absence of a sieving matrix. Follow-up studies (5, 6) have attributed the aggregation to an electrohydrodynamic instability triggered by a coupling of macroion (DNA) concentration fluctuations and electric field induced flows beyond the Debye length scale in a moderately concentrated DNA solution (around the DNA overlap concentration c à ). These results shed light onto why it is so difficult to separate large DNA molecules using capillary electrophoresis.Over the last decade there has been a shift from techniques measuring ensemble-averaged molecular properties (e.g., the aforementioned capillary electrophoresis experiments), to techniques that manipulate single, dilute DNA molecules in micro/ nanofluidic devices or nanopores (7-9). Electric fields are a convenient mode to transport DNA as they scale favorably with device dimensions. The long standing belief in such dilute, single molecule experiments is that uniform DC (direct current) electric fields (up to a few hundred V∕cm in standard electrophoresis buffers) do not greatly perturb the conformation of large DNA unless some sort of sieving matrix is used (e.g., gel or microfabricated post array) (1). Numerous fluorescence microscopy experiments confirm this belief to be true [see (10) for a recent review], but are typically limited to field strengths of a few tens of V∕cm to avoid image blur. Much larger electric fields give rise to some chain orientation and possibly some slight chain stretching (11), though the stretching is inconclusive because it is not directly meas...

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