Thermostable DNA polymerase for high accuracy
Thermus species, recombinant, E. coli
For in vitro use only!
Unit Definition: One unit is defined as the amount of the enzyme required to catalyze the incorporation of 10 nmol of dNTP into an acid-insoluble form in 30 minutes at 74 °C.
Shipping: shipped on blue ice
Storage Conditions: store at -20 °C
avoid freeze/thaw cycles
Shelf Life: 12 months
Concentration: 2.5 units/μl
High Fidelity Pol is based on a blend of Taq DNA polymerase and a proofreading enzyme specially designed for highly accurate and efficient amplification. It shows excellent results with extremely long (up to 30 kb), GC-rich or other difficult templates.
The enzyme blend includes a highly processive 5'→3' DNA polymerase and possesses a 5'→3' polymerization-dependent exonuclease replacement activity. Its inherent 3'→5' exonuclease proofreading activity results in a greatly increased fidelity of DNA synthesis compared to Taq polymerase.
The enzyme is highly purified and free of bacterial DNA.
Fidelity of the enzyme:
High Fidelity Pol is characterized by a 4-fold higher fidelity compared to Taq polymerase.
ERHigh Fidelity Pol = 3.4 x 10-6
The error rate (ER) of a PCR reaction is calculated using the equation ER = MF/(bp x d), where MF is the mutation frequency, bp is the number of base pairs of the fragment and d is the number of doublings
(2d = amount of product / amount of template).
High Fidelity Pol (red cap)
2.5 units/μl High Fidelity Polymerase in storage buffer
High Fidelity Buffer (green cap)
Recommended 50 μl PCR assay:
|5 μl||10x High Fidelity Buffer||green cap|
|200 μM||each dNTP||-|
|0.2 - 0.5 μM||each Primer||-|
|1 - 100 ng||template DNA||-|
|High Fidelity Pol||red cap|
|Fill up to 50 μl||PCR-grade water||-|
|95 °C||2 min||1x|
|denaturation||95 °C||20 sec||20-30x|
|annealing1)||50 - 68 °C||30 sec||20-30x|
|elongation2)||68 °C||1 min/kb||20-30x|
|68 °C||1 min/kb||1x|
For optimal specificity and amplification an individual optimization of the recommended parameters may be necessary for each new template DNA and/or primer pair.
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