>>I would like to open a discussion on DNA melting temperature - in particular
>with regard to oligonucleotides. I would like to design primers for PCR, and
>I am currently using the Breslauer nearest neighbour values of entropy and
>enthalpy to calculate the Tm based on the nucleotide sequence.
>>HOWEVER, what if there is a mismatch? (obviously this is not desirable for
>designing primers but it is useful to know if it binds anywhere else...) In
>this case, how is the Tm calculated? I have seen, in Manatius, the suggestion
>that 1 degree C should be subtracted for each mismatch (?) but this is based
>on the Suggs equation ( Tm = 4 * (GC) + 2 * (AT) etc.) !
>>>Stuff deleted
>--
>Dorothy Lowry
>Dept. Medical Biochemistry
>University of Geneva, Switzerland dlowry at cmu.unige.ch>Graduate student and PC/GENE programmer phone: +41 22 70.25.490
>>I may be a bit naive about this, but it seems to me that the only way to
obtain the exact Tm of your missmatched oligos you should just do the
experiment (taking advantage of the hypochromicity effect).
I think that when you take into consideration all the factors involved in the
Tm of any sequence (stacking interactions, deformation of the backbone due to
missmatch, etc...) you realize that you'll never be able to predict it with a
simple algorithm.
Cheers.
_________________________________________________________________
Michael van Waes |
Div. of Biol. Sci. | "I never worry about the future,
Univ. of Montana | ... it comes soon enough. "
Phone: 406-243-5733 |
bi__mvw at selway.umt.edu | Albert Einstein.
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