Melting temperature

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The dissociation of a double-stranded DNA molecule is often referred to as melting because it occurs quickly once a certain temperature has been reached. For multiple copies of a specific DNA molecule (usually synthesized by polymerase chain reaction), the melting temperature (T_m) is defined as the temperature at which 50% of that same DNA molecule species form a stable double helix and the other 50% have been separated to single strand molecules. The melting temperature depends on both the length of the molecule, and the specific nucleotide sequence composition of that molecule.

1 Methods
- 1.1 Wallace method
2 References
3 See also
4 External links

[edit] Methods

Nearest-neighbor model provides the most accurate predictions (average error less than 2^oC). Several sets of thermodynamic parameters and formulas were reported to calculate T_m values<ref>SantaLucia J.,Jr. and Hicks D. (2004) The thermodynamics of DNA structural motifs. Annu. Rev. Biophys. Biomol. Struct. 33, 415-440.</ref><ref> Breslauer, K.J. et al. (1986) Proc. Natl. Acad. Sci. USA. 83, 3746-3750. (pdf) </ref><ref> Rychlik, W. et al. (1990) Nucleic Acids Res. 18, 6409-6412. </ref><ref>Owczarzy R., You Y., Moreira B.G., Manthey J.A., Huang L., Behlke M.A., and Walder J.A. (2004) Effects of sodium ions on DNA duplex oligomers: improved predictions of melting temperatures. Biochemistry 43, 3537-3554.</ref>. Some formulas are more accurate in predicting melting temperatures of DNA duplexes<ref>Owczarzy R., Vallone P.M., Gallo F.J., Paner T.M., Lane M.J. and Benight A.S. (1997) Predicting sequence-dependent melting stability of short duplex DNA oligomers. Biopolymers 44, 217-239. (pdf)</ref>.

[edit] Wallace method

The fastest and less accurate (average error more than 10^oC) Wallace method was used in the past for oligos less than 18mers in length by counting the frequency of each nucleotide base.

The determination of T_m for DNA extracted from organisms was originally designed to determine the GC content of the DNA using the UV absorbance profile as a function of temperature <ref> Mandel, M. and J. Marmur (1968) Methods in Enzymology 12 Nucleic Acids B, 195-206. For long chain DNA polymers CsCl density gradient analysis </ref><ref> Meselson, M. F. W. Stahl and J. Vinograd (1957) Proc. Natl. Acad. Sci. USA. 43, 581-588 yields values which correlate with the melting temperatures </ref><ref> Mandel, M. C., Schildkraut and J. Marmur (1968) Methods in Enzymology 12 Nucleic Acids B: 184-195. Where failures to correlate the buoyant density and the melting temperature arise (as in many bacteriophage DNAs) unusual bases have been found replacing all or part of one of the nucleotides in the genome.</ref>.