Frequently Asked Questions About Our Products And Services

For your convenience, please use our Oligo Analysis Tool.

The extinction coefficient at 260 nm (e₂₆₀) is a unique physical property of each oligonucleotide. It is defined as the absorbance at 260 nm of a 1 M aqueous solution measured at 20 °C in an optical cell with 1 cm pathway (Lambert-Beer's law).
Purinic bases show a higher absorption (OD₂₆₀) than pyrimidinic bases. Interactions between neighbouring bases, as well as modifications that absorb at 260nm, also influence optical absorbance. As a consequence, the extinction coefficient strongly depends on oligonucleotide sequence and composition.

The formula what Eurofins has always used to calculate the amount of substance is:
nmol = OD x 100 / (1.54 x nA + 0.75 x nC + 1.17 x nG + 0.92 x nT)
(n = number of respective nucleotides)

Now, Eurofins has also implemented a new formula to make the calculation of the molar extinction coefficient and thus the determination of amount of substance more accurate. The new formula is based on the nearest-neighbor method and takes into account the sequence specific effects.

nmol = OD x 10⁶ / ε260  

Nearest-neighbor and individual-bases extinction coefficients are used in the nearest-neighbors formula when calculating the final, total extinction coefficient of an oligonucleotide:

For your oligos you can decide in your personal preferences which formula should be used to calculate the oligo concentration.

For your convenience, please use our Oligo Analysis Tool or apply the formula below:

N [nmol] = OD x 100 / 1.54 x n_A + 0.75 x n_C + 1.17 x n_G + 0.92 x n_T

n = number of nucleosides of the type X


Example:
Sequence: GAA ATG AGT GCT CAT CAC TAC TTC CGC (27mer)

n_A=7; n_C=8; n_G=5; n_T=7
OD=12 (measured)

N [nmol] = 12 x 100 / 1.54 x 7 + 0.75 x 8 + 1.17 x 5 + 0.92 x 7
= 41.3

For your convenience, please use our Oligo Analysis Tool or apply the formula below:

M[µg] = N x MW / 1000
MW = Molecular weight


Example:
Sequence: GAA ATG AGT GCT CAT CAC TAC TTC CGC (27mer)

n_A=7; n_C=8; n_G=5; n_T=7
MW (calculated) = 8219 g/mol
N= 41.3 nmol

M[µg] = 41.3 x 8219 / 1000 = 339

For your convenience, please use our Oligo Analysis Tool or apply the formula below:

MW [g/mol]= 249.23 x n_A + 240.21 x n_T + 265.23 x n_G + 225.20 x n_C + 63.98 x (s-1) + 2.02

n = number of nucleosides of the type X
s = number of all nucleosides per sequence


Example:
Sequence: GAA ATG AGT GCT CAT CAC TAC TTC CGC (27mer)

n_A=7; n_C=8; n_G=5; n_T=7

MW [g/mol]= 249.23 x 7 + 240.21 x 7 + 265.23 x 5 + 225.20 x 8+ 63.98 x (s-1) + 2.02 = 8219.33

The melting temperature T_M, characterises the stability of the DNA hybrid formed between an oligonucleotide and its complementary strand.

For your convenience, please use our Oligo Analysis Tool or apply the formula below:

Sequences with 15 or less bases 
T_M [°C] = 2(n_A + n_T) + 4(n_G + n_C)

Sequences with more than 15 bases
T_M [°C] = 69.3 + [41(n_G + n_C) / s - (650 / s)]

n = number of nucleosides of type X 
s = number of all nucleosides per sequence 


Example:
Sequence: GAA ATG AGT GCT CAT CAC TAC TTC CGC (27mer)

nA=7; nC=8; nG=5; nT=7 s= 27

Sequences with 15 or less bases
T_M [°C] = 2(7 + 7) + 4(5 + 8) = 80.0

Sequences with more than 15 bases
T_M [°C] = 69.3 + [41(5 + 8) / 27] - (650 / 27) = 69.3 + 19.7 - 24.0 = 65.0

PCR                  T_A [°C] = [(T_M1 + T_M2) / 2] - 3

Sequencing        T_A [°C] = T_M + 3