UV-Melting Curves

UV-Melting Points

One traditional method to determine the stability of DNA or RNA duplexes is to heat an aqueous solution of the duplex in question and observing the change in the UV absorption of the solution as the temperature increases. Upon dissociation of the duplex, the base stacking is lost, so that the UV absorption increases. Plotting the UV absorption against the temperature then yields a so-called melting curve. The temperature at which half of the duplex is dissociated is called the 'melting point'. The more stable the duplex, the higher the UV-melting point. Standard settings are a heating rate of 1 °C/min and UV monitoring at 260 nm. From the UV melting curves, enthalpy and entropy of duplex formation/dissociation can be derived.

Reference

Breslauer, K.J. Extracting thermodynamic data from equilibrium melting curves for oligonucleotide order-disorder transitions. Methods Enzymol. 1995, 259, 221-242.