TopTaq DNA Polymerase

Read phase-out notice below
For highly reliable end-point PCR applications with unrivaled ease of use

Features

  • Storage at 4°C allows immediate reaction setup
  • Convenient room-temperature handling
  • A single, preoptimized protocol suitable for all PCR assays
  • High yields of specifically amplified DNA

Product Details

TopTaq DNA Polymerase combines a number of convenient and unique features to provide unrivaled ease of use and ensure high DNA yields in a wide range of PCR applications. The unique TopTaq Stabilizer included in the enzyme storage buffer allows storage of TopTaq DNA Polymerase at 4°C and reaction setup at room temperature without time-consuming thawing of reagents. CoralLoad Concentrate, which contains two gel-tracking dyes for easier pipetting visualization and immediate gel-loading of PCR products, is also provided. High yields of PCR products are achieved, even when amplifying a range of different sized products using the same Mg2+ concentrations and annealing temperatures.

 

IMPORTANT NOTE: This product is phasing-out until October 15, 2020, and afterward, available only until stocks last. Visit the product page of the successor kit to view improved features or to request a trial kit.

 

For more information and FAQs on this transition, visit: www.qiagen.com/PCRresource.

Performance

TopTaq DNA Polymerase is used for standard and specialized end-point PCR applications. TopTaq DNA Polymerase is the first PCR kit that can be stored at 4°C. TopTaq Stabilizer (contained in the enzyme storage buffer) binds to TopTaq DNA Polymerase, preventing polymerase inactivation during long-term storage. During the initial denaturation step, the TopTaq Stabilizer dissociates from the polymerase without compromising polymerase activity. TopTaq DNA Polymerase outperformed DNA polymerases tested from other suppliers and ensures high yields of specific products, even when using low template amounts, and independent of fragment size (see figure " High PCR product yields without optimization).
See figures

Principle

TopTaq DNA Polymerase is a recombinant 94 kDa thermostable DNA Polymerase. TopTaq DNA Polymerase has been developed by QIAGEN to provide highly reliable end-point PCR with unrivaled ease of use. Until the development of the TopTaq PCR system, all PCR enzymes required storage at –20°C; however, due to the unique proprietary TopTaq Stabilizer contained in the enzyme storage buffer, TopTaq DNA Polymerase is the first PCR kit that can be stored at 4°C. 

High-yield amplification

TopTaq DNA Polymerase, in combination with the innovative TopTaq PCR Buffer, facilitates amplification of specific PCR products by providing a high ratio of specific-to-nonspecific primer binding. Owing to a uniquely balanced combination of KCl and (NH4)2SO4, the PCR buffer provides stringent primer-annealing conditions over a wider range of annealing temperatures and Mg2+ concentrations than conventional PCR buffers (see figure " Increased specificity of primer annealing"). The need to optimize PCR by varying the annealing temperature or the Mg2+ concentration is dramatically reduced, or often not required. Thus, TopTaq DNA Polymerase provides the same reliability and specificity as Taq DNA Polymerase and additionally offers improved performance in some PCR systems — resulting in higher yields of PCR products (see figure " High yields of PCR product without optimization").

TopTaq Stabilizer

TopTaq Stabilizer binds to TopTaq DNA Polymerase at 4°C and room temperature, preventing polymerase denaturation during long-term storage. Template DNA and primers are also prevented from binding to the polymerase at low temperature. During the initial denaturation step, the TopTaq Stabilizer dissociates from the polymerase without compromising polymerase activity (see figure " Stabilization at 4°C").

CoralLoad Concentrate

For enhanced convenience, TopTaq DNA Polymerase is supplied with CoralLoad Concentrate. CoralLoad Concentrate contains a gel-loading reagent and two gel-tracking dyes. This unique reagent improves pipetting visibility and facilitates estimation of DNA migration distance and optimization of agarose gel run time. When using CoralLoad Concentrate, PCR products can be directly loaded onto an agarose gel without prior addition of loading buffer (see figure " Convenient CoralLoad Concentrate").

Optimized protocol and buffer

With its balanced potassium and ammonium ions, the unique PCR Buffer, used in combination with TopTaq DNA Polymerase, promotes  specific primer–template annealing and simultaneously reduces nonspecific annealing. Maximum yields of specific products are obtained even when using low template amounts.The unique buffer formulation and single preoptimized protocol eliminate the need for optimization of experimental parameters for individual-primer template systems, saving time and costs. High yields of specific amplification product are achieved, even when amplifying a range of fragments of different sizes using the same Mg2+ concentration and annealing temperature (see figure " High PCR product yields without optimization").

Q-Solution

This innovative PCR additive facilitates amplification of difficult templates by modifying the melting behavior of DNA. Use of this unique reagent will often enable or improve suboptimal PCR. Unlike DMSO and other PCR additives, Q-Solution is used at a defined working concentration with any primer-template system and is not toxic.

See figures

Procedure

Due to the unique proprietary TopTaq Stabilizer contained in the enzyme storage buffer, TopTaq DNA Polymerase is the first PCR kit that can be stored routinely at 4°C. During the initial denaturation step, the TopTaq Stabilizer freely dissociates from the polymerase without compromising polymerase activity. This added convenience allows immediate reaction setup without the use of ice — saving the time usually required for reagent thawing (see flowchart " TopTaq procedure"). All components can be combined  at room temperature eliminating the need for working on ice. The unique buffer formulation and a single preoptimized protocol eliminate the need for optimization of  experimental parameters for individual primer–template systems.

See figures

Applications

TopTaq DNA Polymerase is highly suited to all end-point PCR applications.

Supporting data and figures

Specifications

FeaturesSpecifications
applicationsPCR, RT-PCR, DNA fingerprinting
dntpsincludedNo
enzymeactivity5' -> 3' exonuclease activity
realtimeorendpointEndpoint
reactiontypePCR amplification
mastermixNo
sampletargettypeGenomic DNA and cDNA
singleormultiplexSingle
storageconditionsFridge storage and room temperature setup
withwithouthotstartWithout Hotstart

Resources

Kit Handbooks (1)
For standard and specialized end-point PCR applications without the need for optimization
Brochures & Guides (4)

FAQ

Which nucleotide analogs can be used with TopTaq DNA Polymerase?

Nucleotide analogs that can be used with TopTaq DNA Polymerase are:

  • dNTP
  • ddNTP
  • biotin-dATP
  • dITP
FAQ ID -1741
Is TopTaq DNA Polymerase a hot-start polymerase?

No. TopTaq DNA Polymerase is not a hot-start polymerase. Although TopTaq Stabilizer reduces enzyme accessibility at lower temperatures, it does not allow stringent hot-start procedures.

 

FAQ ID -1748
Does TopTaq Stabilizer of the TopTaq DNA Polymerase and Master Mix Kits interfere with downstream applications?

No. TopTaq Stabilizer contained in the enzyme storage buffer of the TopTaq DNA Polymerase and TopTaq Master Mix Kits does not interfere with downstream applications such as cloning, sequencing, or restriction digestion.

 

 

FAQ ID -1744
Do CoralLoad dyes supplied in various QIAGEN PCR Kits interfere with downstream applications?

CoralLoad dyes supplied in PCR Kits such as, e.g., Taq, HotStarTaq, and TopTaq DNA Polymerase and TopTaq Master Mix do not interfere with most downstream enzymatic applications.

However, for reproducible results, purification of PCR products using the QIAquick or MinElute PCR Purification Kits prior to enzymatic manipulation is recommended.

 

 

FAQ ID -1745
How comparable is CoralLoad gel loading dye contained in various QIAGEN PCR Kits to Sigma Red?

CoralLoad gel tracking dye contained in Taq, HotStarTaq, TopTaq DNA Polymerase and TopTaq Master Mix Kits separates into 2 fragment-size dependent colors (orange and red) when loaded onto an agarose gel. Sigma Red buffer only has one color which is harder to visualize.

 

 

FAQ ID -1644
What is the shelf-life of TopTaq DNA Polymerase and Master Mix?

TopTaq DNA Polymerase and the TopTaq Master Mix Kit are shipped on dry ice but retain full activity at room temperature (15-25°C) for at least 2 weeks. Both kits can be stored at 2-8°C for up to 12 months.

 

 

 

FAQ ID -1743
What is the upper length of PCR products that can be amplified with TopTaq DNA Polymerase and Master Mix Kits?

The maximum amplicon length we have tested using TopTaq DNA Polymerase and the TopTaq Master Mix Kit was 5 kb, amplified from genomic DNA template. Larger PCR amplicon sizes are possible when using less complex template DNA.

 

 

 

 

FAQ ID -1738
What is the fidelity of TopTaq DNA Polymerase?

The error rate of TopTaq DNA Polymerase is very similar to that of standard Taq DNA Polymerase: approximately 2-3x 10e-5 (per base, per cycle).

 

 

 

 

FAQ ID -1739
Can TA/UA cloning be done with PCR products amplified with TopTaq DNA Polmyerase and TopTaq Master Mix Kits?

Yes. The terminal transferase activity of TopTaq DNA Polymerase adds a 3`A-overhang to the PCR product and therefore enables TA- and UA- cloning.

 

 

 

FAQ ID -1740