Cr. Sabanayagam et G. Holzwarth, REAL-TIME VELOCITY OF DNA BANDS DURING FIELD-INVERSION GEL-ELECTROPHORESIS, Electrophoresis, 17(6), 1996, pp. 1052-1059
The velocity v of bands of double-stranded, linear DNAs containing 48.
5-5700 kbp was determined with 0.3s resolution during field-inversion
agarose gel electrophoresis (FIGE) for a broad range of the forward pu
lse period T-+, keeping the duration of the backward pulse T = T-+/3.
Within 0.6 s or less after the field changed sign from - to +, the vel
ocity showed a sharp positive peak; a similar spike, but with negative
velocity, occurred immediately after the field changed from + to -. F
or long pulses, the magnitude of this spike increased with M(0.36), re
aching ten times the steady-state velocity for M = 5.7 kbp. After this
spike, the velocity dipped to 55-75% of its value in a steady field,
then increased to a small secondary peak before reaching a steady-stat
e plateau. The duration of the velocity trough, and the time of the sm
all peak, increased as M(1). For standard FIGE conditions (ratio of fo
rward:reverse pulse duration, T-+:T = 3:1; equal forward and reverse f
ield amplitudes, E(+) = E(-)), the mobility mu = integral vdt over a c
omplete cycle was a minimum when E(+) terminated at the end of the vel
ocity trough. The minimum occurred because the velocity during E(+) sa
mpled primarily the trough, and because the backward velocity during E
(-) was exceptionally large; the negative velocity spike was maximized
when T-+ terminated at the end of the velocity trough. Computer simul
ations of FIGE by Zimm (J. Chem. Phys. 1991, 94, 2187-2206) and by Duk
e and Viovy (J. Chem. Phys. 1992, 96, 8552-8563) generate real-time ve
locities that are in excellent agreement with our experimental data.