Single Molecule Measurements of Trapped Circular DNA During Electrophoresis in Agarose Gels
Kenneth D. Cole, Adolfas K. Gaigalas, B. Akerman
The effect of agarose gel concentration on the electrophoretic trapping of open (relaxed) circular DNA was investigated using microscopic measurements of individual molecules stained with a fluorescent dye. Three open circles with sizes of 52.5 kilo basepairs (kbp), 115 kbp, and 220 kbp were trapped by the electric field (6 V/cm) and found to be predominately fixed at a single point in the gel and stretched. The length of the stretched circles did not significantly change with agarose concentration of the gels (mass fractions of 0.0025, 0.01, and 0.02). The relaxation kinetics of the trapped circles was measured in the gels. The relaxation of the large open circles was found to be a slow process, taking a number of seconds. The velocity and average length of the 52.5 kbp open circles and 48.5 kbp linear DNA were measured during electrophoresis in the agarose gels. The velocity increased when the agarose concentrations were lowered, but the average length of the open circle DNA (during electrophoresis) did not significantly change in the different agarose gel concentrations. The circles displayed the characteristic pattern of cyclic stretching and relaxation during electrophoresis similar to linear DNA.
agarose gel, electrophoresis, open circle DNA, relaxation, trapping