Melting Temperature (Tm) Calculation
Source code is freely downloable at biophp.org
Basic Melting Temperature (Tm) CalculationsTwo standard approximation calculations are used.
When degenerated nucleotides are included in the primer sequence (Y,R,W,S,K,M,D,V,H,B or N), those nucleotides will be internally substituted prior to minimum and maximum Tm calculation.
Example: Primer sequence: CTCTRYCTWSCTCTCT Sequence for minimum Tm calculation: CTCTATCTAGCTCTCT Sequence for maximum Tm calculation: CTCTGCCTAGCTCTCT
Equations above assume that the annealing occurs under the standard conditions of 50 nM primer, 50 mM Na+, and pH 7.0.