The velocity and recoil of DNA bands during gel electrophoresis

Abstract

The velocity and recoil of bands of DNA containing 48.5 to 4800 kilobasepairs (kb) were measured during pulsed‐field gelelectrophoresis by a video imaging and analysis system. When a 10 V/cm electric field was first applied, the velocity showed an initial sharp peak after approximately 1 s whose amplitude depended on the molecular weight of the DNA and the rest time and polarity of the previous pulse. For example, G DNA (670 kb) exhibited an initial velocity peak of 13 μm/s. The velocity then oscillated through a shallow minimum and small maximum before reaching a 5.0 μm/s plateau. After the field was turned off, the bands moved backward (recoiled). The band position obeyed a stretched‐exponential relation, x = x 0 exp[ − (t/τ)β] with amplitude x 0 equal to approximately 1/10th of the DNA contour length and β≊0.6; for S. pombeDNA,x 0 was a remarkable 165 μm. Both the initial velocity spike and the recoil arise from the presence of a significant fraction of U‐shaped molecules with low configurational entropy. The initial velocity spike is exploited in field‐inversion gelelectrophoresis to generate the ‘‘antiresonance,’’ which is the basis of size‐dependent mobility. Recent computer simulations which include tube‐length fluctuations and tube leakage are in excellent accord with the measuredvelocities.