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HotStarTaq Master Mix Kit

For highly specific amplification for any PCR application

  • High PCR specificity without the need for optimization
  • Easy reaction setup at room temperature
  • Ready-to-use master mix format reduces pipetting steps
HotStarTaq Master Mix contains HotStarTaq DNA Polymerase, the unique QIAGEN PCR Buffer that minimizes the requirement for optimization, and dNTPs. Providing all components in a master mix reduces pipetting steps and the risk of contamination, while increasing throughput and reproducibility.
Cat No./ID: 203443
HotStarTaq Master Mix Kit (250 U)
$259.00
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3 x 0.85 ml HotStarTaq Master Mix (contains 250 units HotStarTaq DNA Polymerase, PCR Buffer with 3 mM MgCl2, and 400 µM of each dNTP)and 2 x 1.7 ml RNase-Free Water
Cat No./ID: 203445
HotStarTaq Master Mix Kit (1000 U)
$915.00
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12 x 0.85 ml HotStarTaq Master Mix (contains 1000 units HotStarTaq DNA Polymerase, PCR Buffer with 3 mM MgCl2, and 400 µM of each dNTP) and 8 x 1.7 ml RNase-Free Water
Cat No./ID: 203446
Hot StarTaq Master Mix Kit (2500 U)
$2,074.00
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1 x 25 ml HotStarTaq Master Mix (contains 2500 units HotStarTaq DNA Polymerase PCR Buffer with 3 mM MgCl2, and 400 µM of each dNTP) and 1 x 50 ml RNase-Free Water
The HotStarTaq Master Mix Kit is intended for molecular biology applications. This product is not intended for the diagnosis, prevention, or treatment of a disease.
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Specific amplification in multiplex PCR.
Fragments from the murine p53 gene were amplified from genomic DNA in multiplex PCR. Parallel reactions were prepared using standard reaction conditions and an enzyme from Supplier R (No hot start) or using HotStarTaq Master Mix Kit from QIAGEN (HotStarTaq Master Mix). M: markers.
Performance

Each lot of HotStarTaq Master Mix Kit 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 Master Mix Kit 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 temperatures and magnesium concentrations").

The combination of high specificity and easy handling makes the HotStarTaq Master Mix Kit suitable for use with complex genomic or cDNA templates (see figure "Effect of hot start on RT-PCR performance"), multiple primer pairs (see figure "Specific amplification in multiplex PCR"), and templates isolated from difficult sources or very low-copy targets (see figure "Highly sensitive single-cell PCR"). It is also suitable for projects such as genetic screening, in which large numbers of samples are amplified.

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 Master Mix is a ready-to-use mixture of HotStarTaq DNA Polymerase, QIAGEN PCR Buffer, and dNTPs. HotStarTaq DNA Polymerase, a modified form of Taq DNA Polymerase, provides high specificity in hot-start PCR.

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 figures "Wide annealing temperature window" and "Tolerance to variable magnesium concentration").

Procedure
HotStarTaq Master Mix Kit is supplied in a convenient master mix format for maximum ease of use. HotStarTaq DNA Polymerase is activated by a 15-minute, 95°C incubation step, which can easily be incorporated into existing thermal cycling programs. Room-temperature reaction setup using the master mix is fast and easy — simply pipet 25 µl HotStarTaq Master Mix into each PCR tube and add 25 µl of primers and template DNA diluted in the RNase-free water provided with the kit (see figure "HotStarTaq procedure"). Pipetting steps are minimized, reducing the possibility of errors and contamination, and ensuring increased throughput and reproducibility. The kit includes a streamlined, optimized protocol for fast and easy PCR setup.
Applications

HotStarTaq Master Mix Kit is highly 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 Yes
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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User-Developed Protocols (1)
As starting material, 5 g soil was mixed with different amounts of Bacillus subtilis cells. Sensitivity was 5 x 103 cells/5g soil.
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References
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