(468c) The Electrophoretic Migration of Partially Denatured dsDNA in a Gel: Why Does It Block?

Gel electrophoresis can separate partially denatured dsDNA fragments based on their chemical sequence. For example, upon an increase in temperature, AT-rich regions melt into two strands before GC-rich regions do. This melting changes the conformations of the DNA molecules during electrophoresis and thus affects the mobility. In practice, the mobility of partially melted DNA molecules is severely reduced, and small differences in composition often lead to large differences in mobility. In spite of the tremendous power of separation of the methods based on this well-known phenomenon, we still do not know the relation between the mobility and the conformations of the partially melted DNA molecules. The only available model, which is several decades old, predicts that the reduction in mobility is directly proportional to the average number of denatured bases regardless of their positions along the DNA chain. In this presentation, we will first derive this old model using De Gennes’ reptation ideas, and we will show that the relevant parameters are not what the literature claims. We will then show how Langevin Dynamics simulations clearly demonstrate that the position of the melted regions is as important as their size, in contradiction with the assumptions made by the previous models. In fact, our results indicate that melted regions at the ends of the molecules have much more impact than melted regions (bubbles) between the two ends. Finally, again using simulations, we identify squid-like molecular conformations as being responsible for the drop in mobility upon melting. Our study also suggests new ways to optimize these methods.