Enzymes

Enzymes

Choosing the Enzymes for PCR can profoundly affect the outcome of the PCR. As PCR became more sophisticated, PCR Enzymes such as polymerase mixtures and blends began to be used.

The primary requirements for a DNA polymerase used in PCR are optimal activity at temperatures around 75°C and the ability to retain that activity after prolonged incubation at even higher temperatures (95°C). The first thermostable DNA polymerase to be widely used for PCR was Taq DNA Polymerase. For many conventional PCRs that do not require extensive optimization, Taq DNA Polymerase is still a good choice. High quality, such as recombinant Taq DNA Polymerase produces the best results. Nevertheless a major drawback of standard Taq DNA Polymerase is its activity at temperatures below its optimum of 72°C. In non-optimized systems, this will lead to formation of primer-dimers due to elongation of primers annealed to each other before the first DNA denaturation step has occurred. Minerva Biolabs PCR Enzymes such as Taq DNA Polymerase is a highly processive 5´→3´ polymerase with no 3´→5´ exonuclease (proofreading) activity. At 72 °C and around pH 9 conditions, MB Taq DNA Polymerase reaches its highest level of activity, capable of amplifying a standard 1 kb strand of DNA in approximately 30 seconds. The Taq‘s activity is blocked at ambient temperature, e.g. while preparing your sample. The inhibition of the Taq polymerase is completely reversed when the temperature is above 70 °C. Complete activation of the polymerase is achieved at 94 °C for 2 min.

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