TAQ DNA POLYMERASES

ORDERING INFORMATION

TAQ DNA POLYMERASE (5 U/µl)

Catalog #	Units	10x Reaction Buffer	50 mM MgCl2	Price (EUR)
# Taq 100	100	1 x 1.8 ml	1.5 ml	14,90
# Taq 500	500	2 x 1.8 ml	1.5 ml	57,90

RED TAQ DNA POLYMERASE (1 U/µl)

Catalog #	Units	10x Reaction Buffer	50 mM MgCl2	Price (EUR)
# Red 100	100	1 x 1.8 ml	1.5 ml	16,80
# Red 500	500	2 x 1.8 ml	1.5 ml	65,50

GREEN TAQ DNA POLYMERASE (5 U/µl)

Catalog #	Units	5x Reaction Buffer	50 mM MgCl2	Price (EUR)
# Green 100	100	2 x 1.8 ml	1.5 ml	16,80
# Green 500	500	4 x 1.8 ml	1.5 ml	65,50

DESCRIPTION

Taq DNA Polymerase is a highly purified recombinant thermostable DNA polymerase that has been isolated from E. coli carrying a vector encoding the Thermus aquaticus DNA polymerase gene. The enzyme possesses a highly processive 5’→3’ DNA polymerase activity with optimum activity achieved at 74°C.
It exhibits high thermal stability in withstanding prolonged incubations at elevated temperatures (95°C). Taq DNA Polymerase lacks 3’→5’ exonuclease activity.

Red Taq DNA Polymerase contains an inert red dye which allows identification of the reactions which contain enzyme. The dye has no adverse effect on PCR.

Green reaction buffer contains two dyes (blue and yellow) that separate during electrophoresis to monitor migration progress. Reactions assembled with Green reaction buffer have sufficient density for direct loading onto agarose gels. Green reaction buffer is recommended for any amplification reaction that will be visualized by agarose gel electrophoresis and ethidium bromide staining. The dyes absorb between 225–300nm, making standard A260 readings to determine DNA concentration unreliable.

Unit Definition

Enzyme Storage Buffer

10x Reaction Buffer

Mg²⁺-Solution

Endodeoxyribonuclease Assay

Exodeoxyribonuclease Assay

No detectable degradation of lambda DNA/HindIII fragments was observed after incubation of 10 units of Taq DNA Polymerase with 1 µg of digested DNA resp. 1 µl of digested DNA in 50 µl of Taq Buffer with KCL containing 1.5 mM MgCl₂ for 4 hours at 37°C resp. 70°C.

Ribonuclease Assay

Functional Assay

BASIC PROTOCOL

Mix the following components on ice in a thin-walled 0.2 ml PCR-tube:

Component	Volume (in µL)	Final concentration
10x Reaction Buffer	5	1x
50 mM MgCl2 solution	1.5	1.5 mM
12.5 mM dNTP-Mastermix	1	250 µM
Forward-Primer	variable	0.2 – 1 µM
Reverse-Primer	variable	0.2 – 1 µM
Template DNA	variable	< 1 µg
Red Taq DNA Polymerase (1 U/µl)	1.0 – 2.5	1 – 2.5 U
Taq DNA Polymerase (5 U/µl)	0.2 – 0.5	1 – 2.5 U
2x distilled, sterile water	Add to a final volume of 50
Total volume	50

CYCLING PROGRAM

Step	Temperature (in °C)	Time	Cycles
Initial activation	94	5 min	1
Denaturation	94	30 sec	23 – 35
Annealing	53*	45 sec	23 – 35
Extension	72	30 sec/kb	23 – 35
Final Extension	72	30 sec/kb	1

* : Approximately 5°C below T_m of primers

HINTS AND NOTES

Reaction conditions (incubation temperatures and times, concentrations of template DNA, primers, magnesium ions and enzyme) depend on template and primers used. Optimal MgCl₂ concentrations vary between 1 – 4 mM and have to be determined empirically. However, many applications work at the standard concentration of 1.5 mM MgCl₂. Advanced applications on genomic DNA require higher MgCl₂ concentrations (2 – 3 mM) adjustable with the separate 50 mM MgCl₂ solution supplied with the set.

OPTIMIZATION OF MgCl₂ CONCENTRATION IN REACTION MIXTURE

Final concentration of MgCl2 in 50 µl reaction volume	Add 50 mM MgCl2 solution
1.5 mM	1.5 µl
1.75 mM	1.75 µl
2.0 mM	2.0 µl
2.5 mM	2.5 µl
3.0 mM	3.0 µl
4.0 mM	4.0 µl

STORAGE

Store at -20°C for 24 months.

WARNING