Single-Molecule Kinetics Show DNA Pyrimidine Content Strongly Affects RNA:DNA and TNA:DNA Heteroduplex Dissociation...

The heteroduplex hybridization thermodynamics of DNA with either RNA or TNA are greatly affected by DNA pyrimidine content, where increased DNA pyrimidine content leads to significantly increased duplex stability. Little is known, however, about the effect that purine or pyrimidine content has on the hybridization kinetics of these duplexes. In this work, single-molecule imaging is used to measure the hybridization kinetics of oligonucleotides having varying DNA pyrimidine content with complementary DNA, RNA, and TNA sequences. Results suggest that the change in duplex stability from DNA pyrimidine content (corresponding to purine content in the complementary TNA or RNA) is primarily due to changes in the dissociation rate, and not single-strand ordering or other structural changes that increase the association rate. Decreases in heteroduplex hybridization rates with pyrimidine content are similar for RNA and TNA, indicating that TNA behaves as a kinetic analogue for RNA.