The required thermal induction of this enzyme makes reaction setup faster and easier than when using Amplitaq DNA Polymerase by eliminating the conventional hot-start step of heating the reaction prior to the addition of the enzyme. Amplitaq Gold DNA Polymerase has been shown to enhance multiplex reactions, especially those with marginal primer balance or suboptimal conditions, by increasing the specificity of all desired targets. It also serves as a tracking dye in gel samples, migrating a little faster than bromophenol blue. PCR was performed on five different primer-template sets using 5 units of Taq DNA polymerase alone or with the addition of 5 units of Taq Extender PCR additive. Image provided by Stratagene.
Enzyme selection can be an important factor when dealing with difficult templates.
However, the high fidelity proofreading enzymes generally require careful optimization to give good yields. This step will likely require optimization, particularly in multiplex assays if the individual primers display significant differences in melting temperature. If the annealing temperature is too low, unwanted non-specific bands will appear on the gel, and if it is too high, expected bands may be missing. Once again, multiplex reactions may likely require some trial and error to enhance the amplification of fragments less robust than others. Generally, 20-30 cycles are sufficient for many applications.