The acceleration of linear DNA during pulsed-field gel electrophoresis.

ABSTRACT

The velocity and orientation of T4 and lambda DNA have been measured for the first 20 s during pulsed-field gel electrophoresis in order to clarify the DNA motions that occur. For a square pulse with field strength E = 10 V/cm, the velocity of lambda DNA increases gradually to 10.5 microns/s in 1.0 s, declines to 8.6 microns/s, and then rises to a plateau value of 9.3 microns/s after 4 s. T4 DNA behaves similarly, but more slowly. Parallel measurements of fluorescence-detected linear dichroism show that the DNA becomes substantially aligned with its chain axis parallel to the electrophoretic field E after the pulse is applied. The alignment also shows an overshoot, an undershoot, and a plateau comparable to those seen for velocity. When the field strength increases, both the velocity and the alignment reach their peaks more quickly. For all field strengths and both molecular weights, the velocity peak occurs when the molecular center of mass has moved 0.3 to 0.5 L, where L is the chain contour length. A qualitative model is provided.