This tool calculates melting temperature (Tm), molecular weight (MW), Extinction Coefficient (OD/μmol, μg/OD) for oligos
Please input the oligo sequence in the text-box below.
1. Sequence (Paste the raw sequence, not fasta format):
Full length:0
Extinction (absorption) coefficients of nucleic acids calculated at 260 nm
This calculation tools depends on an approach which is based on the nearest-neighbor model and its published parameters for DNA and RNA1,2. These parameters are presented in the table below and were determined at wavelength of 260 nm, and neutral pH. The average error of calculated extinction coefficients was shown to be around 4 % under these conditions.
The table of extinction coefficients [liter/(mol.cm)] for DNA and RNA oligonucleotides.
| DNA | RNA | ||
|---|---|---|---|
| Stack or monomer | Extinction coefficient | Stack or monomer | Extinction coefficient |
| pdA | 15400 | pA | 15400 |
| pdC | 7400 | pC | 7200 |
| pdG | 11500 | pG | 11500 |
| pdT | 8700 | pU | 9900 |
| dApdA | 27400 | ApA | 27400 |
| dApdC | 21200 | ApC | 21000 |
| dApdG | 25000 | ApG | 25000 |
| dApdT | 22800 | ApU | 24000 |
| dCpdA | 21200 | CpA | 21000 |
| dCpdC | 14600 | CpC | 14200 |
| dCpdG | 18000 | CpG | 17800 |
| dCpdT | 15200 | CpU | 16200 |
| dGpdA | 25200 | GpA | 25200 |
| dGpdC | 17600 | GpC | 17400 |
| dGpdG | 21600 | GpG | 21600 |
| dGpdT | 20000 | GpU | 21200 |
| dTpdA | 23400 | UpA | 24600 |
| dTpdC | 16200 | UpC | 17200 |
| dTpdG | 19000 | UpG | 20000 |
| dTpdT | 16800 | UpU | 19600 |
Extinction coefficients are typically used to determine oligonucleotide concentrations from the Lambert-Beer law,
concentration(mol/l) = A260nm/(ε * pathlength(cm))
Melting Temperature (Tm) Calculations
For sequences less than 14 nucleotides the formula is3: Tm = (nA + nT)*2 + (nG + nC)*4
For sequences longer than 13 nucleotides, the equation used is4: Tm= 64.9 +41*(nG + nC - 16.4)/(nA + nT + nG + nC)
where n is the number of the bases A,T,G,C in the sequence, respectively.
Molecular Weight Calculations
DNA Molecular Weight (typically for synthesized DNA oligonucleotides. The DNA Molecular Weight Calculations assume that there is not a 5' monophosphate)
Anhydrous Molecular Weight = (An x 313.21) + (Tn x 304.2) + (Cn x 289.18) + (Gn x 329.21) - 61.96
An, Tn, Cn, and Gn are the number of each respective nucleotide within the polynucleotide. The subtraction of 61.96 gm/mole from the oligonucleotide molecular weight takes into account the removal of HPO2 (63.98) and the addition of two hydrogens (2.02). Alternatively, you could think of this of the removal of a phosphate and the addition of a hydroxyl, since this formula calculates the molecular weight of 5' and 3' hydroxylated oligonucleotides.
References
- 1. Cantor C.R., Warshaw M.M., and Shapiro H. (1970) Oligonucleotide interactions. III. Circular dichroism studies of the conformation of deoxyoligonucleotides, Biopolymers 9, 1059-1077.
- 2. Fasman, G.D. (Ed.) (1975) Handbook of Biochemistry and Molecular Biology, Volume 1: Nucleic Acids, pp 589, 3rd edition, CRC Press.
- 3. MARMUR J, DOTY P. Determination of the base composition of deoxyribonucleic acid from its thermal denaturation temperature. J Mol Biol. 1962
- 4. Wallace,R.B., Shaffer,J., Murphy,R.F., Bonner,J., Hirose,T., and Itakura,K. (1979) Nucleic Acids Res 6:3543-3557