Oligonucleotide Properties Calculator

Calculate core properties for DNA and RNA oligonucleotides: Tm (nearest-neighbor), molecular weight, extinction coefficient, OD260 concentration, GC content, and thermodynamic constants. Use Primer Analyzer when a sequence also needs hairpin, dimer, mismatch, or BLAST specificity review.

Quick Start

Enter SequenceDNA (A,T,C,G) or RNA (A,U,C,G)
Set ParametersType, concentration, salt
CalculateGet all properties instantly
Export ResultsCopy for lab notebook

Enter Oligonucleotide Sequence

Length: 0 nt

OD calculations are for single-stranded DNA or RNA

OD Calculation Parameters

If provided, concentration and micrograms will be calculated from OD260

Modifications (Optional)

Press Ctrl + Enter to calculate

Results

No results yet

Enter a sequence and click "Calculate"

What Is the Oligo Properties Calculator?

The Oligo Properties Calculator is a streamlined tool that computes all essential physical and thermodynamic properties of a DNA or RNA oligonucleotide from its sequence. In a single analysis, it reports: molecular weight, GC content, reverse complement, extinction coefficient (ε260), OD260 conversions (nmol/OD, µg/OD), melting temperature using three methods (Basic, Salt-adjusted, Nearest-neighbor), and full thermodynamic parameters (ΔH, ΔS, ΔG, RlnK).

This quick property sheet is designed for researchers who need compact characterization of an oligonucleotide, for example when receiving oligos from a vendor and preparing stock solutions. For primer review with hairpins, self-dimers, hetero-dimers, BLAST specificity checking, and mismatch effects, use the Primer Analyzer.

The calculations follow published methods including SantaLucia 1998 nearest-neighbor parameters for Tm, Owczarzy salt correction, and sequence-based extinction coefficient methods. Use the outputs as sequence-level estimates and check vendor documentation when modifications or reporting conventions affect the final order.

How to Use the Oligo Properties Calculator

  1. Enter your DNA or RNA sequence in the input field. The calculator accepts standard IUPAC codes.
  2. All properties are calculated instantly — molecular weight, GC%, Tm (3 methods), ε260, and thermodynamic constants.
  3. Use the OD260 input to convert absorbance readings to molar concentration (useful when measuring with NanoDrop).
  4. Copy the reverse complement for designing the paired primer or for ordering the complementary strand.
  5. Review ΔG to assess duplex stability — more negative values indicate stronger binding.

Frequently Asked Questions

What are the thermodynamic parameters ΔH, ΔS, and ΔG?
ΔH (enthalpy change) represents the heat released or absorbed during duplex formation — more negative values indicate stronger hydrogen bonding, typically -200 to -500 kcal/mol for 20-mers. ΔS (entropy change) represents the disorder decrease when two strands form a duplex — typically -500 to -1500 cal/(mol·K). ΔG (Gibbs free energy) combines both: ΔG = ΔH - TΔS. At the melting temperature, ΔG = 0. More negative ΔG at 37°C indicates a more stable duplex.
How is the reverse complement generated?
The reverse complement is created by: (1) complementing each base (A↔T, G↔C for DNA; A↔U, G↔C for RNA), and (2) reversing the resulting sequence to maintain 5'→3' directionality. For example, the reverse complement of 5'-ATCGATCG-3' is 5'-CGATCGAT-3'. IUPAC ambiguity codes are also complemented: R↔Y, S↔S, W↔W, K↔M, B↔V, D↔H, N↔N.
Why are there three different Tm calculation methods?
Basic Tm uses the Wallace Rule (2°C per A/T + 4°C per G/C) — suitable for rough estimates of oligos <14 nt, accuracy ±5°C. Salt-adjusted Tm adds a correction for monovalent cation concentration — accuracy ±3°C. Nearest-neighbor Tm uses thermodynamic stacking parameters for each dinucleotide step — accuracy ±1-2°C. For primer design, always use nearest-neighbor Tm as it accounts for sequence context that the other methods ignore.
What does RlnK represent?
RlnK is the gas constant (R = 1.987 cal/(mol·K)) multiplied by the natural log of the equilibrium constant (K) for the duplex formation reaction. It represents the favorability of duplex formation under standard conditions. RlnK = ΔH/Tm - ΔS. A more negative RlnK indicates a more favorable duplex equilibrium (more molecules in duplex form vs single-stranded form at a given temperature).
Is this tool suitable for modified oligonucleotides?
The calculator provides accurate results for unmodified DNA and RNA sequences. For 5'-phosphorylated oligos, the Molecular Weight Calculator offers explicit phosphate modification support. For other modifications (biotin, fluorophores, LNA, 2'-O-methyl), the standard nearest-neighbor parameters may not be accurate — use the calculated values as a baseline and consult modification-specific literature for Tm corrections. LNA typically raises Tm by 2-6°C per substitution.

Related Tools

Primer Analyzer

Primer review with secondary structure checks, BLAST specificity checking, mismatch effects, and application-specific presets.

Tm Calculator

Dedicated Tm tool with batch processing for analyzing hundreds of sequences simultaneously.

Dilution Calculator

Use MW and ε260 values from this tool to prepare accurate stock solutions.

Review request

Check a result or ordering detail

Send the calculation, settings, or pool submission detail that needs a second look.

Related reading

Continue with the page or tool that matches the next decision in your experiment.

Validate PCR Primers Before OrderingDesign GuideMethod References