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An Introduction to PCR Primer Design and Optimization of Amplification Reactions

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Forensic DNA Profiling Protocols

Part of the book series: Methods in Molecular Biology ((MIMB,volume 98))

Abstract

PCR optimization is usually performed in order to obtain maximum specificity and yield. In some applications for which the amount of template may be limiting, or when there is a large amount of nontarget sequences, the sensitivity is maximized. Nonoptimized conditions promote artifactual bands resulting from primer dimerization and mispriming, broad bands containing a mixture of correct and incomplete products, and, generally speaking, a poor signal-to-noise ratio. Poorly designed PCR protocols also lack robustness. Such situations can occur in which the PCR is challenged with less than pristine DNAs or when the ingredients of the reaction are not added in the exact amounts as described by the authors of the protocol. Thus, a well-optimized PCR should be reliable in the hands of a coworker or in the laboratory of another institute.

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  1. Perkin-Elmer Applied Biosystems, Foster City, CA

    James M. Robertson & Jennifer Walsh-Weller

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  1. James M. Robertson
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  2. Jennifer Walsh-Weller
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Editors and Affiliations

  1. University of London, UK

    Patrick J. Lincoln

  2. LGC, Teddington, UK

    Jim Thomson

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© 1998 Humana Press Inc., Totowa, NJ

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Robertson, J.M., Walsh-Weller, J. (1998). An Introduction to PCR Primer Design and Optimization of Amplification Reactions. In: Lincoln, P.J., Thomson, J. (eds) Forensic DNA Profiling Protocols. Methods in Molecular Biology, vol 98. Humana Press. https://doi.org/10.1385/0-89603-443-7:121

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  • DOI: https://doi.org/10.1385/0-89603-443-7:121

  • Publisher Name: Humana Press

  • Print ISBN: 978-0-89603-443-3

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