Taq DNA Polymerase 2x Master Mix Red
Suggested Protocol using Taq 2x Master Mix
(1.5 mM MgCl2 final concentration)
This protocol serves as a guideline for primer extensions. Optimal
reaction conditions such as incubation times, temperatures, and
amount of template DNA may vary and must be determined
individually.
Cat. No.
Size
Reactions
Taq DNA Polymerase
2x Master Mix Red, 1.5 mm MgCl2
Cap colour
Red
180301
100
2 x 1.25 ml
180303
500
10 x 1.25 ml
180306
2.500
Other product sizes available
50 x 1.25 ml
Storage and Stability: The unopened kit is stable at -20 °C for 2 years
after the production date. Reagent for in vitro laboratory use only.
Key Features





Significantly reduced set up time
Diminished sources of contamination
Increased reproducibility
Facilitates confirmation of complete mixing
Direct loading of products onto agarose gels
Notes:

Set up reaction mixtures in an area separate from that used for
DNA preparation or product analysis. Work on ice at all times.

The table below shows the reaction set up for a final volume of
50 L. If desired, the reaction size may be scaled down.

After primer extension, a sample (10 to 30% of the reaction)
can be loaded directly on a gel for analysis.

Thaw Taq 2x Master Mix and primers. It is important to thaw
the solutions completely and mix thoroughly before use to
avoid localized concentrations of salts. Keep all components
on ice.
1.
Set up each reaction as follows:
General Description
Taq DNA Polymerase 2x Master Mix Red is a ready-to-use 2x reaction
mix. Simply add primers, template, and water to successfully carry
out primer extensions and other molecular biology applications.
+
NH4
Ampliqon Taq DNA Polymerase, the
buffer system, dNTPs and
magnesium chloride are present in Taq DNA Polymerase 2x Master
Mix Red. An inert red dye and a stabilizer are also present to allow
direct loading of the final products onto an agarose gel for analysis.
Each reaction requires 25 µL of the 2x reaction mix. Simply add
primers, template and water to a total reaction volume of 50 µL.
Taq DNA Polymerase 2x Master Mix Red offers several advantages.
Set up time is significantly reduced. There is no need to buy and use
separate loading dyes to load reaction products onto agarose gels for
electrophoresis and subsequent visualization. The chance of
contaminating component stocks is eliminated. Reduction of reagent
handling steps leads to better reproducibility. Standard tests can be
set up with the confidence that results will be consistent every time.
Composition of the Taq DNA Polymerase 2x Master Mix (1.5 mM
MgCl2 final concentration)




Tris-HCl pH 8.5, (NH4)2S04, 3 mM MgCl2, 0.2% Tween 20
0.4 mM dNTPs
0.2 units/µl Ampliqon Taq DNA polymerase
Inert red dye and stabilizer
Component
Vol./reaction
Final Conc.
Taq 2x Master Mix Red
25 µL
1x
Primer A
Variable
0.1 – 1.0 µM
Primer B
Variable
0.1 – 1.0 µM
Distilled Water
Variable
----
Template DNA
Variable
Variable
TOTAL volume
50 µL
----
2.
Mix gently by pipetting the solution up and down a few times.
3.
Program the thermal cycler according to the manufacturer´s
instructions.
4.
For maximum yield and specificity, temperatures and cycling
times should be optimized for each new template target or
primer pair.
5.
Place the tubes in the thermal cycler and start the reaction.
Three-step PCR program
Cycles
Duration of cycle
a
25 - 35
20 – 30 seconds
b
20 – 40 seconds
c
30 seconds
d
1
5 minutes
a.
Temperature
95 °C
50 – 65 °C
72 °C
72 °C
Denaturation step: This step is the first regular cycling event and
consists of heating the reaction to 95 °C for 20 – 30 seconds. It causes
melting of the DNA template by disrupting the hydrogen bonds
between complementary bases, yielding single-stranded DNA
molecules.
b.
c.
d.
Annealing step: The reaction temperature is lowered to 50 – 65 °C for
20 – 40 seconds allowing annealing of the primers to the singlestranded DNA template. Typically, the annealing temperature is about
3 – 5 °C below the Tm of the primers used.
Extension/elongation step: Taq DNA polymerase has its optimum
activity temperature at 72 °C. At this step the DNA polymerase
synthesizes a new DNA strand complementary to the DNA template
strand. The extension time depends on the length of the DNA fragment
to be amplified. As a rule of thumb, at its optimum temperature the
DNA polymerase will polymerize a thousand bases per minute.
Final elongation: This single step is occasionally performed at a
temperature of 72 °C for 5 minutes after the last PCR cycle to ensure
that any remaining single‐stranded DNA is fully extended.
Ampliqon A/S
Stenhuggervej 22
DK-5230 Odense M
Denmark
Phone: +45 70201169
Fax: +45 70201179
info@ampliqon.com
www.ampliqon.com
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Description
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dNTP Mix (2 x 500 µl): 50 mM mixed dNTPs
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(Other product sizes available)
Tween 20 is a registered trademark of ICI Americas, Inc.
Cat. No.
100103
110103
110203
110303
110403
110503
111304
112103
120301
150301
160301
180301
190301
200303
210303
210403
220303
230301
230701
232701
250407
250507
290401
290801
501004
511104
NOTICE
In certain countries, patents cover the PCR process. This product is intended
for researchers having a license to perform PCR or those not required to
obtain a license.
Issued 04/2012 am/jp