We demonstrate that agarose gel electrophoresis of linear duplex DNA in the presence of ethidium bromide has a marked effect on the mobility of genomic DNA fragments detected by Southern hybridization. Mobility shifts of greater than 6% of the actual molecular weight were detected when different amounts of the same DNA sample were analyzed in 1.0% agarose gels containing 0.5 micrograms ml-1 ethidium bromide. For forensic applications, shifts of this magnitude could complicate the task of comparing restriction fragment length polymorphism profiles and introduce a degree of uncertainty to allele frequency population databases.