Color OptiTaq DNA Polymerase (Dyes-traced Proofreading Blend & Long PCR)

Mixture of stable thermophilic DNA polymerases capable of generating PCR products up to 20 kb with high fidelity;, suitable for applications requiring high temperature synthesis of DNA.

The enzyme is supplemented with two inert gel tracking dyes.

Description:

• OptiTaq DNA Polymerase is a modified and optimized thermostable enzymes blend containingThermus aquaticus DNA polymerase and Pyrococcus sp. DNA polymerase.
• Ultrapure, recombinant enzymes are used to prepare OptiTaq DNA Polymerase.
• The enzymes blend exhibits the 3´->5´ proofreading activity, resulting in considerably higher PCR fidelity and processivity than possible with unmodified Taq DNA polymerase (1).
• Enables increased amplification product yield in comparison with Taq DNA polymerase over wide range of PCR products.
• Maintains the 5'´->3´ exonuclease activity.
• Adds extra A at the 3' ends.
• Suitable for multiplex PCR as it exhibits wider tolerance for Mg²⁺, salts concentration and pH (2,3).
• Improves PCR results with critical templates, such as containing GC-rich regions, palindromes or multiple repeats.
• Color OptiTaq 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 20 kb.
• Use of Color OptiTaq 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 min 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 Conditions: Store at -20°C.

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.

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 20 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. Cline, J., Braham, J. and Hogrefe, H. (1996) Nucleic Acids Res. 24, 3545.
2. Chien, A., Edgar, D.B. and Trela, J.M. (1976) J. Bacteriol. 127, 1550.
3. A.S.,Sliusarenko, A.G. and Gorodetskii, S.I. (1980) Biokhimiya 45, 644.