PCR Primer Tm Calculator

PrimerX

Check the Melting Temperature of PCR Primers.

The bioinformatics software, PrimerX, has been designed to calculate the melting temperature (Tm) of a pair of primers based on the nearest neighbour modelling. The nearest neighbour model considers the entropy and enthalpy of nucleotides based upon their specific arrangement in the DNA sequence. These values can then be adjusted for salt and primer concentrations to output a melting temperature. Information about the sequence length and G-C content are also provided.

The melting temperature (Tm) is a measure of the temperature at which the double-stranded DNA structure of a molecule begins to denature and separate into single strands. The Tm is calculated using the following formula: Tm = (1000 x ΔH) / (ΔS + R ln (DNA /2)) - 273.15, where...

ΔH is the enthalpy of melting
ΔS is the entropy of melting
R is the gas constant
DNA is the concentration of the DNA molecule
"1000" is a conversion factor that converts the units of ΔH from calories per mol to joules per mol
"ln" is the natural logarithm function, which is used to calculate the logarithm of the DNA concentration value
"2" is a constant value that is used in the formula to calculate the Tm
"-273.15" is a conversion factor that converts the units of temperature from degrees Celsius to degrees Kelvin.

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