GC Content Melting Temperature Calculator
Calculate DNA melting temperature from GC percentage using Marmur-Schildkraut-Doty equation: Tm = 81.5 + 16.6 log[Na+] + 0.41(GC%) - 600/n. For longer DNA fragments and standard conditions.
🧬 Molecular Biology📐 Tm = 81.5 + 16.6log[Na+] + 0.41(%GC) - 600/n🧬 Biology
GC content (%)
Sequence length (bp)
Na+ concentration (M)
Please enter valid values.
Formula & Reference
| Variable | Symbol | Formula | Units |
|---|---|---|---|
| GC Content Melting Temperature Calculator | — | Tm = 81.5 + 16.6log[Na+] + 0.41(%GC) - 600/n | degrees C |
Step-by-Step Examples
Example 1
Typical Genome
Human genome average GC=41%, n=3e9 (effectively infinite), 50mM Na+.
- Tm = 81.5 + 16.6xlog(0.05) + 0.41x41 - 0
- Tm = 81.5 - 22.0 + 16.8 - 0 = 76.3 degrees C
- Genomic DNA melts broadly due to heterogeneous GC content
- Actual Tm range: 60-90 degrees C across genome
✓ Tm ~ 76 degrees C for human genome average
Example 2
GC-Rich Sequence
GC=70%, n=200 bp, 50mM Na+.
- Tm = 81.5 - 22.0 + 0.41x70 - 600/200
- Tm = 81.5 - 22.0 + 28.7 - 3.0 = 85.2 degrees C
- High GC regions are more thermally stable
✓ Tm = 85.2 degrees C
Example 3
AT-Rich Region
GC=30%, n=150 bp, 1M Na+.
- Tm = 81.5 + 16.6xlog(1) + 0.41x30 - 600/150
- Tm = 81.5 + 0 + 12.3 - 4.0 = 89.8 degrees C
- High salt increases Tm significantly
✓ Tm = 89.8 degrees C at 1M Na+
Real-World Applications
FISH Hybridization
Chromosome painting probes need Tm optimization for stringent hybridization conditions.
DNA Stability Analysis
Predicting thermal denaturation profiles for genomic regions.
Restriction Enzyme Work
Understanding local melting during enzyme recognition and cutting.
Genomics Research
AT-rich vs GC-rich regions have different melting profiles affecting DNA extraction protocols.
Common Mistakes to Avoid
⚠️
Applying long-DNA formula to short oligos
Use Wallace rule (Tm = 2AT + 4GC) for oligos < 20 bp. MSD equation applies to longer DNA where end effects are negligible.
⚠️
Ignoring ion concentration effects
Salt concentration dramatically affects Tm (+16.6 degrees C per 10-fold increase in [Na+]). Always report Tm with salt concentration.
⚠️
Treating Tm as a single value for genomic DNA
Genomic DNA has a broad melting transition due to heterogeneous GC content, not a single Tm.
Frequently Asked Questions
What is DNA melting? ▾
Denaturation of double-stranded DNA to single strands. Driven by heat breaking hydrogen bonds and base stacking. Monitored by increased absorbance at 260nm (hyperchromic effect).
Why does GC content raise Tm? ▾
G-C base pairs form 3 hydrogen bonds, A-T pairs form 2. More G-C pairs = more bonds to break = higher melting temperature.
How does salt affect Tm? ▾
Na+ ions stabilize DNA by neutralizing negatively charged phosphate backbone, reducing electrostatic repulsion between strands. Higher [Na+] = higher Tm.
What is a melting curve? ▾
Plot of absorbance (or fluorescence) vs temperature. DNA melts over a range around Tm. Tm is the midpoint (50% melted). Used in diagnostics (HRM analysis).
What is HRM analysis? ▾
High-Resolution Melting analysis uses precise melting curves to detect SNPs, mutations, or methylation differences. Used in clinical diagnostics.