Abstract
Separation of single-base substitution sequential DNA isomers remains one of the most challenging tasks in DNA separation by capillary electrophoresis. We developed a simple, versatile capillary electrophoresis technique for the separation of single-base sequential isomers of DNA having the same chain length. This technique is based on charge differences resulting from the different protonation (acid dissociation) properties of the four DNA bases. A mixture of 13 single-base sequential isomers of 12-mer single-stranded DNA was separated by using an electrophoretic buffer solution containing 20mM phosphoric acid (pH 2.0) and 8M urea. We demonstrated that our method could separate all possible mutation patterns under identical experimental conditions. In addition, application of our method to the separation of the polymerase chain reaction product of a 68-mer gene fragment and its single-base isomers indicates that in combination with the appropriate genomic DNA extraction techniques, the method can detect single-base gene mutations.
Published Version
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