Taq Pol High Fidelity Hot Start | Taq DNA Polimerase Hot Start Cellco

Taq Pol High Fidelity Hot Start

Ficha técnica

Código	Taq Pol High Fidelity Hot Start
Categoria	PCR Convencional
Marca	Cellco

Descrição Geral

Taq Pol High Fidelity Hot Start | Taq DNA Polimerase Hot Start - Cellco
Blend de DNA polimerase Hot Start de elevada fidelidade

For in vitro use only!

Definição de unidade: One unit is defined as the amount of the enzyme required to catalyze the incorporation of 10 nmoles of dNTPs into an acidinsoluble form in 30 minutes at 74 °C.

Envio: Shipped on blue ice

Condições de armazenamento: Store at -20 °C

Avoid freeze/thaw cycles

Validade: 12 months

Concentração: 2,5 units/μL

Descrição:

High Fidelity Hot Start Pol is based on a blend of Taq DNA polymerase and a proofreading enzyme specially designed for highly accurate and efficient amplification. The additional hotstart function provides improved specificity and sensitivity when amplifying lowcopy-number targets in complex backgrounds or when prolonged room-temperature set up is required. The polymerase activity is blocked at ambient temperature and switched on automatically at the onset of the initial denaturation. The thermal activation prevents the extension of nonspecifically annealed primers and primer-dimer formation at low temperatures during PCR setup. The enzyme shows excellent results with extremely long (up to 20 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.

Activation step:

High Fidelity Hot Start Pol requires no prolonged heating or denaturing step. The polymerase inhibiting antibodies are released at the increased temperature of the initial denaturation.

Fidelidade da Enzima:

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 d (2d = amount of product / amount of template).

Conteúdo:

High Fidelity Hot Start Pol (blue cap)

2.5 units/µl High Fidelity Hot Start Polymerase in storage buffer

High Fidelity Buffer (red cap)

10x conc.

Recommended 50 µl PCR assay

Component	50 μL rxn	Final conc.
10x HF/HS Reaction Buffer	5 µl	1x
dNTP (Mix 10 mM)	1 µl	200 µM
each primer	0,5 - 2,5 µl	0,2 - 0,5 µM
Taq HF/HS Pol (2,5U/ µl)	0,5µl	1,25 U/reaction
DNA template		10 pg - 1 µg
Water, grade PCR	fill up to 50 µl

Please note that it is essential to add the polymerase as last component.

Incubate reactions in a thermal cycler

Recommended cycling conditions:

Initial denaturation	95 °C	2 min	1x
Denaturation	95 °C	20 sec	20-30 cycles
Annealing**	50 - 68 °C	30 sec	20-30 cycles
Elongation***	68 °C	1 min/kbp	20-30 cycles
Final extension (optional)	68 °C	1 – 2 min/kbp	1x
Hold	4 – 8 °C	∞	1x

**The annealing temperature depends on the melting temperature of the primers used

***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.

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