Trapping of megabase-sized DNA molecules during agarose gel electrophoresis

AUTOR(ES)

Gurrieri, Sergio

FONTE

The National Academy of Sciences

RESUMO

Megabase DNA molecules become trapped in agarose gels during electrophoresis if the electric field exceeds a few volts per cm. Fluorescence microscopy reveals that these molecules invariably arrest in U-shaped conformations. The field-vs.-size dependence for trapping indicates that a critical molecular tension is required for trapping. The size of unligated λ-ladders, sheared during gel electrophoresis at a given field, coincides with the size of molecules trapped at that field, suggesting that both processes occur through nick melting near the vertex of the U-shape. Consistently, molecules nicked by exposure to UV radiation trap more readily than unexposed ones. The critical trapping tension at the vertex is estimated to be 15 pN, a force sufficient to melt nicks bent around gel fibers, and, according to our model, trap a molecule. Strategies to reduce molecular tension and avoid trapping are discussed.

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