Melting Temperature (Tm) Calculation

Primer (6-50 bases):
Basic Tm
Degenerated nucleotides are allowed
Base-Stacking Tm
Degenerated nucleotides are NOT allowed
Primer concentration: nM
Salt concentration: mM
Mg2+ concentration: mM

Source code is freely downloable at

Base-Stacking Melting Temperature (Tm) Calculations

This aproximation uses Thermodynamical concepts to compute Tm.

The following references were used to develop the script:

  • SantaLucia J. A unified view of polymer, dumbbell, and oligonucleotide DNA nearest-neighbor thermodynamics. Proc Natl Acad Sci U S A. 1998 Feb 17;95(4):1460-5.NCBI
  • von Ahsen N, Oellerich M, Armstrong VW, Schütz E. Application of a thermodynamic nearest-neighbor model to estimate nucleic acid stability and optimize probe design: prediction of melting points of multiple mutations of apolipoprotein B-3500 and factor V with a hybridization probe genotyping assay on the LightCycler. Clin Chem. 1999 Dec;45(12):2094-101.NCBI