Question: Determination of Tm of non-perfect fitting DNA-duplexes?

Peter J. Floriani, Ph.D. floriani at epix.net
Fri Jan 24 14:28:39 EST 1997


Ming-Piao Kao wrote:
> Does anyone know how one can determine the melting-temperature 
> of 2 DNA-strands which have several mismatches between them ?? 
> And is this possible using GCG software(to which I have access) ?
> Thank you very much for any help,
> De'de' Kao
> mingpiao at sci.kun.nl

When I did my dissertation (in computer science in 1992-1994),
I was assisting biologists who were using this equation:

Given:
   [Na+]         the sodium ion concentration
   percent G+C   the percent G and C in the sequence
   size          the length of the sequence

Compute Tm in degrees centigrade as:

   Tm = 79.8 + 18.5*Log[Na+] + 58.4*(percent G+C) - 820/size

Which was given in:
   Schleicher and Schuell,
   "Transfer and Immobilization of Nucleic Acids 
       to S&S Solid Supports"
   Schleicher and Schuell, Keene, NH, 1987

This source referenced:
   Birnstiel M.L., Sells, B.H., and Purdom, I.F
   "Kinetic Complexity of RNA molecules"
   J. Mol. Biol. 63 (1972) 21-39
and
   Casey J., and Davidson, N.
   "Rates of formation and thermal stabilities of RNA:DNA
      and DNA:DNA duplexes at high concentrations of formamide"
   Nucleic Acids Research, 4 (1977) 1539-1552.   

However, in my dissertation, a "personal communication" (from one 
of the biologists I was assisting) informed me that Tm computed 
with this equation was found to be "insufficient" for predicting 
actual behaviors; in this case, these were probes for in-situ 
hybridization of rRNA.

Without local access to journals, I feel as if I am
giving the melting point of phlogiston rather than DNA and
I expect that this problem has been investigated since then.
So, I would be very interested in hearing from others on 
the computation of Tm.

On the other part of the question: I do not have or use GCG 
and cannot comment on how Tm might be obtained with it.

Peter J. Floriani, Ph.D.
floriani at epix.net

----------------------------------------------------
I deny that biology can destroy the sense of truth, 
which alone can even desire biology. No truth which
I can find can deny that I am seeking the truth.  
My mind cannot find anything which denies my mind. 
G. K. Chesterton, "The Long Bow" in Collected Works 14:96




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