Color Perpetual Taq DNA Polymerase HOT START

Pre-complexed with specific anti-Taq monoclonal antibody, top quality thermophilic Taq DNA polymerase for automatic "hot start" PCR, resulting in greatly enhanced amplification specificity, sensitivity and yield.

Source: Thermus aquaticus

The enzyme is supplemented with two inert gel tracking dyes.

Description:

• Ultrapure, recombinant Taq DNA Polymerase is reversibly complexed with anti-Taq monoclonal antibody that blocks replication activity of the enzyme at moderate temperatures.
• Our carefully selected anti-Taq antibodies have high thermal stability, providing protection against non-specific primer extension from room temperature to 70°C.
• The polymerase activity is restored during the initial denaturation step when amplification reactions are heated at 94-95°C for two minutes.
• Formation of complexes between Taq DNA Polymerase and an anti-Taq antibody forms a basis for automatic "hot start" PCR, which allows for the assembly of PCR reactions at room temperature.
• High stability of the complexes allows for the enormous increase of PCR specificity, sensitivity and yield in comparison to the conventional PCR assembly method.
• Automatic "hot start" PCR provides a fast and convenient method when assembling multiple PCR reactions.
• Both increased specificity and reduced mispriming improve multiplex PCR.
• Eliminated risk of template cross-contamination and assured safe laboratory practice, due to removed necessity to open hot tubes.
• Thermostable Taq Polymerase replicates DNA at 74°C and exhibits a half-life of 40 min at 95°C (1,2).
• Catalyzes the polymerization of nucleotides into duplex DNA in the 5'->3' direction in the presence of magnesium ions.
• Contains the 5'->3' exonuclease activity.
• Lacks the 3'->5' exonuclease activity.
• Adds extra A at the 3' ends.
• Color Perpetual Taq DNA Polymerase is recommended for use in PCR and primer extension reactions at elevated temperatures to obtain a wide range of DNA products up to 10 kb.
• Use of Color Perpetual Taq DNA Polymerase offers several advantages:
  • visualizes the addition of the polymerase to the reaction,
  • confirms complete mixing,
  • enables direct loading of PCR products onto an agarose gel without addition of a gel loading buffer,,
  • the added dyes allow to track electrophoresis progress,
  • do not affect PCR performance,
  • do not interfere with most downstream applications (exeption: the polymerase is not recommended for any downstream application using absorbance or fluorescence excitation).

Unit Definition: One unit is defined as the amount of enzyme required to catalyze the incorporation of 10 nmoles of dNTP into acid-insoluble material in 30 minutes at 74°C. The reaction conditions are: 50 mM Tris-HCl (pH 9.0 at 25°C), 50 mM NaCl, 5 mM MgCl₂, 200 μM each of dATP, dCTP, dGTP, dTTP (a mix of unlabeled and [³H]dTTP), 10 μg activated calf thymus DNA and 0.1 mg/ml BSA in a final volume of 50 μl.

Storage Buffer: 20 mM Tris-HCl (pH 8.0 at 22°C), 100 mM KCl, 0.1 mM EDTA, 1 mM dithiothreitol, 50% glycerol and stabilizers.

Storage Conditions: Store at –20°C.

10 x Reaction Buffer:

10 x Pol Buffer A (optimization buffer without MgCl₂): the buffer allows to optimize MgCl₂concentration.

10 x Pol Buffer B (general application, up to 10 kb): the buffer contains 15 mM MgCl₂ and is optimized for use with 0.2 mM of each dNTP.

Quality Control: All preparations are assayed for contaminating endonuclease, 3'-exonuclease, and nonspecific single- and double-stranded DNase activities. Typical preparations are greater than 95% pure, as judged by SDS polyacrylamide gel electrophoresis.

References:

1. Chien, A., Edgar, D.B. and Trela, J.M. (1976) J. Bacteriol. 127, 1550.
2. Kaledin, A.S., Sliusarenko, A.G. and Gorodetskii, S.I. (1980) Biokhimiya 45, 644.