Comparison of competitively primed and conventional allele-specific nucleic acid amplification

Giffard, Philip M., McMahon, Jennifer A., Gustafson, Helen M., Barnard, Ross T., & Voisey, Joanne (2001) Comparison of competitively primed and conventional allele-specific nucleic acid amplification. Analytical Biochemistry, 292(2), pp. 207-215.

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A simulation of competitively primed allele-specific DNA amplification has been constructed and its behavior examined. This has shown that when the ratio of the amount of homoduplex misprime product to the total amount of amplimer is low, it increases by approximately one-fourth of the mispriming frequency with each doubling of the total amount of amplimer. When the ratio is high and reverse mispriming becomes significant, it asymptotes toward a value <0.5. An analogous simulation was carried out on conventional allele-specific DNA amplification. As expected, the ratio of the amount of amplimer in the positive and negative reactions closely approximates the mispriming frequency provided that amplification is exponential in both cases. This suggests that conventional allele-specific amplification has somewhat higher inherent specificity than competitively primed amplification. However, conventional allele-specific reactions are subject to a "catch-up" phase in which the positive reaction slows or stops, thus reducing the specificity. It was hypothesized that competitively primed reactions may be easier to optimize than conventional allele-specific reactions. This conjecture was supported experimentally. In addition, it was shown that the specificity of competitively primed reactions is a function of the degree of amplification.

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8 citations in Web of Science®
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ID Code: 8474
Item Type: Journal Article
Refereed: Yes
Additional Information: For more information, please refer to the journal's website (see hypertext link) or contact the author.
Author contact details:
DOI: 10.1006/abio.2001.5077
ISSN: 0003-2697
Divisions: Current > Research Centres > CRC for Diagnostics
Past > QUT Faculties & Divisions > Faculty of Science and Technology
Current > Institutes > Institute of Health and Biomedical Innovation
Copyright Owner: Copyright 2001 Elsevier
Deposited On: 06 Jul 2007 00:00
Last Modified: 09 Jun 2010 12:42

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