DNA Melting Temperature Calculator Formula

Understand the math behind the dna melting temperature calculator. Each variable explained with a worked example.

Use the DNA Melting Temperature Calculator

Formulas Used

Tm (Wallace Rule, short oligos)

tm_wallace = 2 * at_count + 4 * gc_count

Tm (Marmur-Doty, long DNA)

tm_marmur = 64.9 + 41 * (gc_count - 16.4) / length

GC Content

gc_content = gc_count / length * 100

Variables

VariableDescriptionDefault
lengthOligonucleotide Length(bp)20
gc_countNumber of G+C Bases10
at_countDerived value= length - gc_countcalculated

How It Works

DNA Melting Temperature

The melting temperature (Tm) is the temperature at which 50% of double-stranded DNA molecules are denatured into single strands.

Wallace Rule (for short oligos < 20 bp)

Tm = 2(A+T) + 4(G+C) °C

Marmur-Doty (for longer DNA)

Tm = 64.9 + 41 × (G+C - 16.4) / N °C

GC base pairs have three hydrogen bonds (vs two for AT), making them more thermally stable.

Worked Example

A 20-mer with 10 GC bases and 10 AT bases.

length = 20gc_count = 10
  1. 01AT count = 20 - 10 = 10
  2. 02Wallace: Tm = 2(10) + 4(10) = 20 + 40 = 60°C
  3. 03GC content = 10/20 × 100 = 50%

Ready to run the numbers?

Open DNA Melting Temperature Calculator