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Oligo Calculation Tool

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

2. Select the modifications: 5'-       3'-



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