Gel electrophoresis of partially denatured DNA. Retardation effect: its analysis and application

The hypothesis about the role of partial denaturatlon in DNA retardation during its electrophoreeis in denaturing gel/1,/2 was tested. We used partially melted DNA molecules in which the size of the melted regions and their location were known. They were obtained through glyoxal treatment of the melted regions by a prooedure allowing the denatured state to be fixed at any point within the melting range. The approach and the availability of the melting maps of DNAs made it possible to investigate DNA molecules differing in length and in the size of the melted regions. The presence of a denatured region at the end of the molecule or inside of it was shown to decrease its electrophoretio mobility, the effect depending on the size of the melted region and on the DNA length. On the basis of the experimental results an explanation is proposed for the cause of retardation in the case of partially denatured DNA.