[Bioperl-l] Bio::SeqFeature::Primer Calculating the Primer TM

john herbert john.herbert at clinical-pharmacology.oxford.ac.uk
Tue Feb 17 04:07:44 EST 2004

Hello Rob,
Thanks for the reply. I agree about Primer3 documentation, had me
puzzled for a while before I sussed it.

At the moment I don't have the code but I guess the easiest way could
be to look at the source code of Primer3 and see what the C code looks
like (I am assuming it is C) for calculating the Primer TM. Then try to
mimic it in Perl. If I get time soon, I will have a look. Alternatively
we could look at the references.

If there are TM Gurus  out there who have a better suggestion, please
mail us. 

Kind regards,


>>> Rob Edwards <redwards at utmem.edu> 17/02/2004 03:40:40 >>>
I wrote this implementation of the Bio::SeqFeature::Primer module, and

I am guilty as charged. I was confused by the primer3 documentation, 
but hey, at least I was honest in the docs - I said that I couldn't 
figure out what was going on.

I don't know what the formula is that Primer3 used. If you have some 
code for this I can update the module.


On Feb 12, 2004, at 5:50 AM, john herbert wrote:

> Hello BioPerl.
> I was thinking of using BioPerl to calculate the Tm of my primers. I
> looked from the documentation and found the method for getting a
> primer's Tm. my $tm = $primer->Tm;
> In the notes of this method, the author is confused by the fact that
> BioPerl calculated Tm never matches that of Primer3's prediction.
> is because Primer3 uses a different method to calculate a Primers TM
> than the method used in the BioPerl.
>  BioPerl $primer->Tm = Calculated using: Tm = 81.5 +
> + .41*(%GC) - 600/length
> (Sambrook, Fritsch and Maniatis, Molecular Cloning, p 11.46 (1989,
> Press).
> Primer3 primer TM = Primer3 uses the oligo melting temperature
> given in Rychlik, Spencer and Rhoads, Nucleic Acids Research, vol
> num 12, pp 6409-6412 and Breslauer, Frank, Bloeker and Marky, Proc.
> Natl. Acad. Sci. USA, vol 83, pp 3746-3750. This method uses
> thermodynamics to calc TM.
> Primer3 does use the Sambrook method to predict the TM of the PCR
> products but not the Primer TM.
> Would it be possible to update the BioPerl to match how Primer3
> calculates the Primer TM?
> Kind regards,
> John Herbert
> Cancer Research UK.
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