How do I measure Tm of the synthesized oligo?
Tm (melting temperature) refers to the temperature where 50% of oligonucleotides exist in duplex form and the rest in single-strand form. There are several ways to calculate Tm. At Bioneer, we use the nearest-neighbor method (PNAS 83, 3746-50). It is believed that the effect of hybridization is different for every sequence and that through thermodynamic measurements; you can estimate Tm values more accurately. For example, the sequences of 5′-GC-3′ and 5′-CG-3′ are different in thermodynamic measurements. The method for nearest-neighbor calculation is as follows: Enthalpy (dH) X 1000 Tm = —————————- – 273.15 + (12.0 X log([Salt])) Entropy(dS) + (R X ln([Oligo])) Through thermodynamic measurements, enthalpy and entropy values are determined between 2 bases. [Salt] is the concentration of monovalent cations and [Oligo] is the oligo concentration. R is the gas constant (1.987 calK-1mole-1). Bioneers Tm value is calculated by the nearest-neighbor method with 50mM for salt
