Calculating Tm - Help Needed

Mayumi Yagi myagi at u.washington.edu
Mon Jul 17 13:36:51 EST 1995


The version I have differs from yours in that the factor is 0.41(percent 
G+C), not fraction G+C.  This would change your calculation to 81.5 -21.6 
+ 20.5 - 20 = 60.4.

On 15 Jul 1995, Shiao Y. Wang wrote:

> 
> I'm using the equation in Sambrook et al. 1989 (pg 11.46) to calculate the
> Tm of oligonucleotides. My results don't look right. Can someone spot my
> mistake?
> 
> The equation is: Tm = 81.5 + 16.6(log[Na+]) + 0.41(fraction G+C) - (600/N)
> Where N = chain length.
> 
> My oligos are 20mers. If Na+ conc is 0.05 M and the oligo is 50% G+C, I get
> 
> Tm = 81.5 + 16.6(log 0.05) + 0.41(0.5) - (600/20) or
> Tm = 81.5 + 16.6(-1.301) + 0.205 - 30 or
> Tm = 81.5 - 21.6 + 0.205 - 30 or
> Tm = 30.1
> 
> I've been doing PCR using an annealing temp of 60C so the 30.1 appears to
> be in error. Thank you very much for any help.
> 
> Shiao Wang
> University of Southern Mississippi
> sywang at whale.st.usm.edu
> 
> 
> 



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