Transition characteristics and thermodynamic analysis of DNA duplex formation: a quantitative consideration for the extent of duplex association

Abstract

Transition characteristics and thermodynamic properties of the single-stranded self-transition and the double-stranded association were investigated and analyzed for 9-, 15- and 21-bp non-self-complementary DNA sequences. The multiple transition processes for the single-stranded self-transition and the double-stranded association were further put forth. The experimental results confirmed that the double-stranded association transition was generally imperfect and the thermodynamic properties of the single-stranded self-transition would exert an influence on a duplex formation. Combining ultraviolet melting experiments in various molar ratios, the extent of duplex association was estimated for three double-stranded DNAs. In our experimental range, the extent of duplex association decreases with increasing the number of base pairs in DNA sequences, which suggest that the short oligonucleotides may proceed in a two-state transition while the long oligonucleotides may not. When the extent of duplex association was considered, the true transition enthalpies of a duplex formation derived from UV and differential scanning calorimetry measurements were in good agreement.

Figure 1

UV relative absorbance versus temperature curves for three dsDNAs in two adding modes and various molar ratios. Column I indicates that S2, S4 and S6 are guest strands and added into host strands S1, S3 and S5, respectively. Column II shows an inverse-adding mode to Column I. Plots for various molar ratios of 20:1 (squares), 10:1 (circles), 5:1 (triangles), 2.5:1 (diamonds) and 1:1 (crosses).

Figure 2

Plots of 1/h_m versus 1/[1 + (m – 1)x_i] (i = A or B) for three duplex association transitions. The experiments were performed in two adding modes: S1 plus various S2 (closed squares), S2 plus various S1 (open squares), S3 plus various S4 (closed circles), S4 plus various S3 (open circles), S5 plus various S6 (closed triangles), S6 plus various S5 (open triangles). It is worth noting that linear fitting does not include the experimental data of the equimolar mixing of two single strands.

Figure 3

Calorimetric excess heat capacity, ΔC_p, versus temperature profiles for the three dsDNAs. Plots for duplex strand transition of S1S2 (squares), S3S4 (circles) and S5S6 (triangles). The thermodynamic parameters derived from these curves are summarized in Table 3.