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Nucleic Acids Research, 2000, Vol. 28, No. 12 E69-e69
© 2000 Oxford University Press

Position and degree of mismatches and the mobility of DNA heteroduplexes

David A. Upchurch, Raj Shankarappa and James I. Mullins*

Department of Microbiology, University of Washington, Seattle, WA 98195-7740, USA

Heteroduplex mobility assay (HMA) is a fast and inexpensive method for determining relatedness between DNA sequences. Rapidly evolving viruses such as HIV-1 develop marked sequence differences in their genomes over the course of the epidemic and infection in a single individual. HMA can be used to monitor both processes. Here, we systematically evaluated the influence of single base mismatches on heteroduplex mobility. The impact of mismatches at nine different positions in 559 bp double-stranded DNA molecules, within a background of overall sequence divergence ranging from 1.97 to 9.65%, was evaluated in both non-denaturing and partially-denaturing acrylamide gels. We found that the electro­phoretic mobility of heteroduplexes was proportional to the level of mismatch when that level exceeded 4.5%. Overall, mismatches near the center of the fragment and clustered mismatches tended to have an exaggerated influence on the mobility of heteroduplexes. Thus, the use of HMA for quantitative inference of genetic distances under the conditions we describe is of greatest utility at levels of mismatch >5%.

* To whom correspondence should be addressed. Tel: +1 206 616 1851; Fax: +1 206 616 1575; Email: jmullins@u.washington.edu


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