Velocity of DNA in gels during field inversion.

Abstract

The velocity and alignment of T4 and \ensuremath{\lambda} DNA have been measured during field-inversion gel electrophoresis (FIGE) at field strengths E between 6 and 14 V/cm. Immediately after E changed sign, the velocity exhibited an oscillatory behavior before reaching a steady value. The times at which these oscillations occurred were shorter for \ensuremath{\lambda} DNA (48.5 kilo-base-pairs) and longer for smaller E. However, the distance each DNA had traveled, when scaled by its contour length L, was about 0.1 for the first velocity peak and 0.15--0.3 for the undershoot for both molecular weights and all values of E studied, provided that E was on for at least 12 s prior to the field inversion. If the field was on for only a short time before inversion, the first sharp peak in velocity was progressively reduced in magnitude and duration. The alignment function f=(3〈${\mathrm{cos}}^{2}$\ensuremath{\theta}〉-1)/2, where \ensuremath{\theta} is the angle between the helix axis and E, was measured for the same DNA's, field strengths, and pulse times. The alignment was at all times parallel to E but decreased sharply immediately after field inversion to a minimum, followed by an overshoot and then a plateau. The time after field inversion at which the minimum in f occurred was approximately the same as that of the velocity maximum. The anticorrelation between the velocity and f, as well as the invariance of x/L for the velocity overshoot, support the idea, first suggested by computer simulations, that DNA undergoing gel electrophoresis spends much time hooked over gel fibers in ssU-shaped chain configurations. The measured velocity curves of T4 DNA were integrated to compute the net displacement after a series of positive and negative field inversions. The average velocity computed from this displacement showed a pronounced ``antiresonance'' or minimum at the same pulse period as observed in practical FIGE separations.