High Fidelity Polymerase
Detailní popis
Thermus species, recombinant, E. coli
For general laboratory use.
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 gel packs
Storage Conditions: store at -20 °C
avoid freeze/thaw cycles
Shelf Life: 12 months
Form: liquid
Concentration: 2.5 units/μl
Description:
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).
Content:
High Fidelity Pol (red cap)
2.5 units/μl High Fidelity Polymerase in storage buffer
High Fidelity Buffer (green cap)
10x conc.
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 | - |
0.5 μl (1.25 units) |
High Fidelity Pol | red cap |
Fill up to 50 μl | PCR-grade water | - |
Please note that it is essential to add the polymerase as last component.
Recommended cycling conditions:
initial denaturation |
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 |
final elongation |
68 °C | 1 min/kb | 1x |
1)The annealing temperature depends on the melting temperature of the primers used.
2)The elongation time depends on the length of the fragments to be amplified. A time of 1 min/kb is recommended.
For optimal specificity and amplification an individual optimization of the recommended parameters may be necessary for each new template DNA and/or primer pair.
Related products:
- Ready-to-Use Mixes / direct gel loading
- Ready-to-Use Mixes
- Thermophilic Polymerases
- Deoxynucleotides (dNTPs)
- Supplements
- Primers and Oligonucleotides
- DNA Ladders
BIOZ Product Citations:
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