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HotStarTaq DNA Polymerase

For highly specific amplification with minimal optimization
  • Minimal optimization requirements
  • High PCR specificity
  • Easy handling and room-temperature setup
HotStarTaq DNA Polymerase uses a chemically mediated hot start that, unlike, antibody-mediated systems, leads to complete inactivation of the polymerase until the initial heat activation step at the start of PCR. HotStarTaq DNA Polymerase is supplied with the unique QIAGEN PCR Buffer, which minimizes nonspecific amplification products, primer dimers, and background. Q-Solution, a novel additive that enables efficient amplification of "difficult" (e.g., GC rich) templates, is also provided.
Cat No./ID: 203203
HotStarTaq DNA Polymerase (250 U)
€143.00
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250 units HotStarTaq DNA Polymerase, 10x PCR Buffer, 5x Q-Solution, 25 mM MgCl2
Cat No./ID: 203205
HotStarTaq DNA Polymerase (1000 U)
€477.00
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4 x 250 units HotStarTaq DNA Polymerase, 10x PCR Buffer, 5x Q-Solution, 25 mM MgCl2
Cat No./ID: 203207
HotStarTaq DNA Polymerase (5000 U)
€2,040.00
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1 x 5000 units HotStarTaq DNA Polymerase, 1 x HotStarTaq Buffer Set (1 x 22 ml PCR Buffer, 1 x 40 ml Q-Solution, 1 x 22 ml MgCl2)
Cat No./ID: 203209
HotStarTaq DNA Polymerase (25000)
€9,002.00
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100 x 250 units HotStarTaq DNA Polymerase, 100 x 1.2 ml HotStarTaq Buffer Set, 100 x 2.0 ml Q-Solution, 100 x 1.2 ml MgCl2
The HotStarTaq DNA Polymerase is intended for molecular biology applications. This product is not intended for the diagnosis, prevention, or treatment of a disease.
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HotStarTaq procedure.

The HotStarTaq procedure is fast and easy for maximum convenience.

1
Superior performance.
A 497 bp fragment was amplified from 50 copies of an HIV-pol-gene construct which had been added to 1 µg human genomic DNA. Different hot-start enzymes were employed: HotStarTaq DNA Polymerase from QIAGEN (HotStarTaq); hot-start enzyme from Supplier AII (Hot-start enzyme); Taq-antibody mixture from Supplier L (Antibody-mediated); enzyme without hot start from Supplier R (No hot start). Specific PCR product is indicated by the arrow. Equal volumes of the reaction were analyzed on a 2% agarose gel. M: markers.
3
Higher specificity with different primer–template systems.
Three different primer–template systems were amplified under the same conditions with either Taq DNA polymerase from Supplier R (R) or with HotStarTaq DNA Polymerase (H). System 1: A 1.1 kb fragment of a D-IgI homolog was amplified from human genomic DNA. System 2: A 296 bp fragment from the chromosomal region correlated with X-linked juvenile retinoschisis was amplified from human genomic DNA. System 3: A 214 bp fragment of the β-actin gene was amplified from cDNA synthesized from total RNA. M: markers.
4
Effect of hot start on RT-PCR performance.
A 1.1 kb fragment of the human interleukin 1 receptor (type II) gene was amplified from cDNA. Amplification reactions were prepared in triplicate using Taq DNA polymerase and buffer from Supplier L(No hot start); antibody-mediated hot start using enzyme and buffer from Supplier L (Antibody-mediated); and HotStarTaq DNA Polymerase and PCR Buffer from QIAGEN (HotStarTaq). M: markers.
5
Highly sensitive single-cell PCR.
A 500 bp fragment of the murine p53 gene was amplified from single cells isolated by flow cytometry, and directly sorted into individual PCR tubes. Reactions were prepared in parallel using HotStarTaq DNA Polymerase and PCR Buffer from QIAGEN (HotStarTaq), a hot-start enzyme and buffer from Supplier AII (Hot-start enzyme), or antibody-mediated hot start and buffer from Supplier L (Antibody-mediated). M: markers.
6
Tolerance to variable temperature and magnesium concentrations.
[A] PCR amplification at the indicated annealing temperatures using QIAGEN PCR Buffer and Taq DNA Polymerase (QIAGEN). The same PCR was performed in parallel using PCR buffer and Taq DNA polymerase from another supplier (Supplier AII). The single-copy human cystic fibrosis gene was amplified. M: markers. [B] Tolerance to Variable Magnesium Concentration. PCR amplification at the indicated Mg2+ concentrations using QIAGEN PCR Buffer and Taq DNA Polymerase (QIAGEN). The same PCR was performed in parallel using PCR buffer and Taq DNA polymerase from another supplier (Supplier AII). The single-copy human prion protein gene was amplified. M: markers.
Performance

Each lot of HotStarTaq DNA Polymerase is subjected to a comprehensive range of quality control tests, including a stringent PCR specificity and reproducibility assay in which low-copy targets are amplified. HotStarTaq DNA Polymerase outperformed kits tested from other suppliers and ensures high specificity and superior performance in hot-start PCR (see figures "Higher specificity with different primer–template systems" and "Superior performance" and table). The innovative PCR buffer provided with the kit ensures specificity over a wide range of PCR conditions, minimizing the need for optimization (see figure Tolerance to variable temperature and magnesium concentrations). Suboptimal PCR can be improved with Q-Solution, also provided with the kit (see figure "Amplification of difficult templates"). Together, these components ensure specific amplification in a range of applications (see figure "Effect of hot start on RT-PCR performance" and "Highly sensitive single-cell PCR").

Comparison of hot-start methods 
HotStarTaq DNA Polymerase Hot-start enzyme from Supplier AII Antibody-mediated Manual Wax barrier
Specific amplification ++ + + +/– +/–
Minimal PCR optimization ++ +/– +/–
Easy to use ++ ++ +
HotStarTaq DNA Polymerase specifications

Concentration: 5 units/µl
Recombinant enzyme: Yes
Substrate analogs: dNTP, ddNTP, dUTP, biotin-11-dUTP, DIG-11-dUTP, fluorescent-dNTP/ddNTP
Extension rate: 2–4 kb/min at 72°C
Half-life: 10 min at 97°C ; 60 min at 94°C
Amplification efficiency: ≥105 fold
5'–>3' exonuclease activity: Yes
Extra A addition: Yes
3'–>5' exonuclease activity: No
Contaminating nucleases: No
Contaminating RNases: No
Contaminating proteases: No
Self-priming activity: No 

  

Principle

HotStarTaq DNA Polymerase, a modified form of Taq DNA Polymerase, provides high specificity in hot-start PCR. The kit includes an innovative dual-cation PCR buffer, Q-Solution, and MgCl2.

HotStarTaq DNA Polymerase

HotStarTaq DNA Polymerase is supplied in an inactive state and has no polymerase activity at ambient temperatures. This prevents extension of nonspecifically annealed primers and primer dimers formed at low temperatures during PCR setup and the initial PCR cycle (see figures "Superior performance in hot-start PCR" and "Higher specificity with different primer–template systems"). HotStarTaq DNA Polymerase is activated by a 15-minute incubation at 95°C, which can be incorporated into any existing thermal-cycler program.

QIAGEN PCR Buffer

QIAGEN PCR Buffer maintains specific amplification in every cycle of PCR by promoting a high ratio of specific-to-nonspecific primer binding during the annealing step in each PCR cycle (see figure "Increased specificity of primer annealing"). Owing to a uniquely balanced combination of KCl and (NH4)2SO4, the buffer provides stringent primer-annealing conditions over a wider range of annealing temperatures and Mg2+ concentrations than conventional PCR buffers. Optimization of PCR by varying the annealing temperature or the Mg2+ concentration is therefore often minimal or not required (see figure Tolerance to variable temperature and magnesium concentrations).  

Q-Solution

Q-Solution, an innovative PCR additive that facilitates amplification of difficult templates by modifying the melting behavior of DNA, is also provided with HotStarTaq DNA Polymerase. This unique reagent improves suboptimal PCR caused by templates that have a high degree of secondary structure or with GC-rich templates (see figure "Amplification of difficult templates"). Unlike other commonly used PCR additives such as DMSO, Q-Solution is used at just one working concentration, is nontoxic, and PCR purity is guaranteed. Adding Q-Solution to the PCR does not compromise PCR fidelity.

Procedure
HotStarTaq DNA Polymerase is supplied with a streamlined, optimized protocol for fast and easy PCR setup. HotStarTaq DNA Polymerase is activated by a 15-minute, 95°C incubation step, which can easily be incorporated into existing thermal cycling programs. Reactions can be set up at room temperature, ensuring greater convenience and ease of use (see figure "HotStarTaq procedure").
Applications

HotStarTaq DNA Polymerase is suitable for a wide variety of applications, including challenging applications, such as amplification of:

  • Complex genomic templates
  • Complex cDNA templates (e.g., RT-PCR)
  • Very low-copy targets (e.g., single-cell PCR)
  • Reactions with multiple primer pairs
Features
Specifications
Applications PCR, RT-PCR, Complex genomic templates, very low-copy targets
Enzyme activity 5' -> 3' exonuclease activity
Mastermix No
Reaction type PCR amplification
Real-time or endpoint Endpoint
Sample/target type Genomic DNA and cDNA
Single or multiplex Single
With/without hotstart With hotstart

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Kit Handbooks (1)
HotStarTaq DNA Polymerase; HotStarTaq Master Mix Kit - For highly specific hot-start PCR without optimization  
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