UV melting analysis

Each oligonucleotide was dissolved in 10 mM sodium phosphate buffer (pH 7.0), 100 mM NaCl, 0.1 mM EDTA to arrange the final concentration of each oligonucleotide to be 2.0 μM. The solution was separated into quarts cells (10 mm) and incubated at 95°C. After 10 min, the solution was cooled to 5°C at a rate of 0.5°C/min. Then, the solution was kept at 5°C for 5 min. After 5 min, the solution was heated at 95°C at the same rate. The absorption at 260 nm was recorded and used to draw UV-melting curves. The obtained melting curves were smoothed using Savitzky–Golay treatment. T_m was calculated as the temperature that gave the maximum of the first derivative of the UV-melting curve.