[Bioperl-l] Bio::SeqFeature::Primer Calculating the Primer TM
jason at cgt.duhs.duke.edu
Tue Feb 17 07:46:25 EST 2004
I believe biopython guys have dealt with this recently -- brad - can you
point us to working code/documentation for Tm calculation?
On Tue, 17 Feb 2004, john herbert wrote:
> 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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jason at cgt.mc.duke.edu
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