4.3. UV Melting Assay

UV-Vis absorption spectra were recorded by a Shimadzu 1700 spectrophotometer (Kyoto, Kyoto, Japan) connected to a thermoprogrammer. 20 μM DNA samples were prepared in a buffer containing 50 mM MES with 0, 100, 500, 1000, 1500 mM KCl in the absence and presence of 40 wt% PEG 200 at pH 6.0. After PEG 200 was added, pH of the buffers were regulated to 6.0, and the concentration of KCl did not affect pH of the solution. We performed UV melting experiments with 20 μM of DNA with 1 or 0.1 cm cell to keep the absorption in a range from 0 to 1.0. After incubating at 90 °C for 3 min, DNA samples were annealed by cooling from 90 °C to 0 °C at a rate of −2 °C·min⁻¹ for triplex formation. The stabilities of triplex DNA were characterized by the changes in UV melting temperatures. A heating rate of 0.5 °C·min⁻¹ was applied in the melting process of DNA samples from 0 °C to 90 °C. The values of T_m were obtained from the peaks of dA/dT plot for the melting curves, and the errors of analysis were ±1 °C.