KlenTaq

For general laboratory use.

Definição de unidade: One unit is defined as the amount of the enzyme required to catalyze the incorporation of 10 nmoles of dNTP's into an acid-insoluble form in 30 minutes at 74 °C.

Envio: shipped on gel packs

Condições de armazenamento: store at -20 °C
avoid freeze/thaw cycles

Validade: 12 months

Forma: liquid

Concentração: 5 units/μl

Descrição:
KlenTaq is a truncated version of Taq DNA Polymerase, lacking its first 280 amino acids. The enzyme catalyzes the polymerization of nucleotides into duplex DNA in 5’→3’ direction in the presence of magnesium but lacks the 5’→3’ exonuclease activity of Taq polymerase. The enzyme is purified by an additional separation process to reduce contaminating bacterial DNA sequences.
In addition to routine PCR, KlenTaq is recommended for genotyping and primer extension. Compared to Taq the enzyme shows an improved fidelity and thermostability.

Contente:
KlenTaq (red cap)
5 units/μl KlenTaq DNA Polymerase in 20 mM Tris-HCl, 100 mM KCl, 0.1 mM EDTA, 1 mM DTT, 0.5 % Tween-20, 0.5 % Nonidet P-40, 50 % (v/v) Glycerol, pH 8.0 (25°C)

KlenTaq Buffer (green cap), 10x conc.
200 mM Tris-HCl, 400 mM KCl, 25 mM MgCl2, pH 8.5 (25 °C)
component||PCR-217S||PCR-217L
KlenTaqPolymerase||200 units/ 40 μl||5 x 200 units/ 5 x 40 μl
KlenTaq Buffer, 10x conc.||400 μl||5 x 400 μl

PCR Reaction Set-Up:
The PCR procedure below shows appropriate volumes for a single 50 μl reaction. For multiple reactions, prepare a master mix of all components and dispense appropriate volumes into each PCR reaction tube prior to adding template DNA and primers.
Thaw up and briefly centrifuge each component before use.
Add the following components to a PCR tube:

Prepare PCR Master Mix

comp.||cap||final conc.||1 assay @ 50 μl
PCR-grade Water||white||||fill up to 50 μl
10x reaction Buffer||green||1x||5 μl
dNTP Mix 10 mM||white||200 μM each||1 μl
KlenTaq||red||2.5 units/reaction||0.5 μl
forward primer(10 μM)||||0.1 - 0.5 μM||0.5 - 2.5 μl
reverse primer(10 μM)||||0.1 - 0.5 μM||0.5 - 2.5 μl
template DNA||||10 pg - 1 μg1)||
1)genomic DNA: 1 ng - 1 μg, plasmid and viral DNA: 1 pg - 1 ng

Mix and briefly centrifuge the components

Optimisation of MgCl2 concentration:
The final assay contains 3.5 mM Mg2+ as recommended for most applications. For an individual optimisation Mg2+ stock solution (#PCR-266) mybe added.

Incubate reactions in a thermal cycler
Recommended cycling conditions:
initialdenaturation||96 °C||2 min||1x
denaturationannealing2)elongation3)||96 °C50 - 68 °C72 °C||15 - 30 sec15 - 30 sec30 sec - 4 min||25 - 35x
final elongation(optional)||72 °C||2 min||1x
hold||4 - 8 °C||||2)The annealing temperature depends on the melting temperature of the primers used.
3)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 optimisation of the recommended parameters may be necessary for each new template DNA and/or primer pair.

Produtos relacionados: Mg2+ Stock, #PCR-266