Electronic ETP-46464 titrations
Electronic spectra of the DNA with the constant concentration of 5.8 × 10−4 M in 0.02 M PBS (pH = 7.4) were recorded in the absence and presence of Ole in two different molar ratios (r = [Ole]/[DNA] = 0.18 and 0.29).
Absorption titration of Ole with the DNA was carried out using a fixed concentration of Ole (5.0 × 10−5 M) in 2.0 mM PBS in the presence of increasing concentrations of the DNA (0, 7.5 × 10−5, 1.5 × 10−4 and 3.0 × 10−4 M).
All experiments were carried out at room temperature. In all experiments, the incubation time for equilibration was 10 min.
Thermal denaturation studies
In trachea studies, 8.0 × 10−5 M dsDNA was thermally denatured into single-strand components in the absence and presence of 2.5 × 10−6 M Ole. At the same time, absorbance of the DNA at 260 nm was monitored at various temperatures. Temperature of the sample was controlled with a thermocouple attached to the sample holder. The temperature was varied from 25 to 95 °C with a constant rate of 2 °C/min and melting temperature (Tm) of DNA was determined as the transition midpoint.