Structural analysis of nucleic acids by precise denaturing gradient gel electrophoresis: II. Applications to the analysis of subtle and drastic mobility changes of oligo- and polynucleotides.

Abstract

Precise denaturing gradient gel electrophoresis was effectively applied to various kinds of oligo- and polynucleotides. The analyses on oligonucleotides revealed that every oligonucleotide has its own characteristic normalized mobility profile (NMP), which can be used to identify, characterize and classify the molecules. The precise system also enabled us to obtain unequivocally the mobility transitions corresponding to the melting of hairpin structures of oligonucleotides, single-stranded (ss) DNAs, and RNAs. Another application to co-migration and separate migration experiments demonstrated that there were significant binding interactions between two species of ss molecules of similar mobility, even when they have little complementarity with each other. When the precise temperature gradient gel electrophoresis was applied to double-stranded DNAs, it could be confirmed with high reliability that the mobility transitions observed correspond to cooperative meltings and strand dissociations. Through these experiments, mu m, a parameter defined as a mobility transition point, was shown to be effective to deal with those phenomena quantitatively.