PerkinElmer LAMBDA 265 - Page 26

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Where, T is the Celsius temperature (°C).
d. Tm method: Select a method for determining Tm (DNA melting temperature). Options
include: 1st derivative and Average.
e. DNA Pair Length (K): Enter the DNA base pair length. If a DNA base pair length is above
5000, enter “0”.
f. Equation Name and Expression: The melting range is calculated within the specified
calculation range by defining the low temperature where the slope begins to increase steadily,
and the high temperature where the slope approaches zero again. The default equation for
the calculation of %G-C (Guanine-Cytosine) base pairs is:
%G-C=2.44 * ( Tm – 81.5 - 16.66 * log(M) + 500/K )
Where, M is the molarity of salt in mol/l, and K is the DNA base pair length.
If a DNA base pair length (K) is entered as ‘0’, then the equation becomes:
%G-C= 2.44 * ( Tm - 81.5- 16.66 * log(M) )
g. Calculation Start Temperature (°C): Enter the start temperature for calculating the Tm value
using the selected method.
h. Calculation End Temperature (°C): Enter the end temperature for calculating the Tm value
using the selected method.
i. Molarity (mol/L): Enter the salt molarity of the solution[mol/l].
4. Select Wavelength Range and enter the wavelength range for the measurement.
5. Select OK in the method dialog box. Select Single Cell Peltier in the Accessory Type on the
main screen of the software.