AffiTaq® Hot Start - With Buffer - With MgCl2 - Without dNTP

Vendor AffiGEN
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AffiTaq® Hot Start - With Buffer - With MgCl2 - Without dNTP

Product Overview:

Description: AffiTaq® Hot Start is an advanced DNA Polymerase formulated with a proprietary buffer solution and MgCl2 for superior PCR performance. This specialized variant features a hot-start technology, minimizing non-specific amplification and primer-dimer formation, ensuring enhanced specificity in PCR reactions. The exclusion of dNTPs allows tailored nucleotide addition based on experimental designs.

Product Components:

Components Concentration AFG-BOR-200 AFG-BOR-201
AffiTaq® Hot Start DNA Polymerase 5 U/μL 1X (200U - 40μL) 5X (1000U - 200μL)
AffiTaq® Buffer - dNTP free & Mg2+ free 10X 1X (40μL) 5X (200μL)
AffiTaq® MgCl2 25mM 1X (40μL) 5X (200μL)

Product Specifications:


  • DNA Polymerase: AffiTaq® sourced from highly purified recombinant enzymes, offering robust and reliable performance, with an added hot-start feature for minimized non-specific amplification.
  • Buffer: A unique proprietary buffer blend optimized for enzyme activity and stability during amplification reactions.
  • MgCl2 Concentration: Included at an optimized concentration for enhanced polymerase activity.

Storage Conditions:

  • Temperature: Store at -20°C for long-term stability and preservation of enzymatic activity.
  • Stability: Maintains stable activity for up to 24 months under recommended conditions.

Performance Characteristics:

Key Features:

  • Hot-Start Technology: AffiTaq® Hot Start minimizes non-specific amplification and primer-dimer formation, ensuring enhanced specificity in PCR reactions.
  • Robustness and Stability: The hot-start variant maintains high robustness and stability in diverse PCR applications.
  • Versatile Usage: Suitable for various PCR applications, including routine PCR, genotyping, and high-throughput PCR.


  • PCR Applications: Ideal for routine PCR, genotyping, and other molecular biology applications requiring high-quality amplification with minimized non-specific products.
  • Sensitivity and Specificity: Demonstrates exceptional sensitivity and specificity across various templates and reaction conditions, improving assay reliability.

Quality Control:

QC Tests Performed:

  • Stringent quality control tests include purity assays, activity assays, and specificity assays, guaranteeing consistent performance and reliability.

Quality Assurance:

  • Manufactured under ISO-certified conditions, maintaining rigorous quality assurance protocols for batch-to-batch consistency and reliability.

Usage Guidelines:

Recommended Protocol:

  • Detailed protocols and guidelines provided for using AffiTaq® Hot Start DNA Polymerase, emphasizing the importance of the initial denaturation step and optimal cycling parameters for superior results.
  • Recommended Concentrations: General recommendations for primer and template concentrations provided for various PCR applications.

Handling Precautions:

  • Adhere to standard laboratory safety practices when handling enzymes. Avoid repeated freeze-thaw cycles for optimal enzyme activity.

Regulatory Compliance:

Certifications and Compliance:

  • Complies with relevant regulatory standards and manufactured in accordance with ISO 9001:2015 guidelines.
  • Certified free of contaminants such as endonucleases, exonucleases, and RNases.

Packaging Information:

Packaging Details:

  • Available in various packaging sizes to suit diverse laboratory requirements, featuring stringent sealing to prevent contamination.
  • Packaging materials ensure stability and integrity during transportation and storage.

Labeling Information:

  • Each product is labeled with batch numbers, expiration dates, storage conditions, and specific composition details for traceability and quality control purposes.

Suggested Protocol for PCR Using AffiTaq® Hot Start - With Buffer - With MgCl2 - Without dNTP

Materials Provided:

  • AffiTaq® Hot Start DNA Polymerase (5 U/μL)
  • AffiTaq® Buffer (10X)
  • AffiTaq® MgCl2 (25mM)

Materials Required but not provided:

  • Primer pairs specific to the target DNA sequence (25)
  • Template DNA
  • Nucleotides (dNTPs) (10 mM)
  • PCR tubes/strips and PCR machine
  • Sterile, nuclease-free water
  • PCR-grade pipettes and tips



  1. Thaw the AffiTaq® Hot Start DNA Polymerase set components on ice.

  2. Prepare a master mix for each PCR reaction as follows:

    • AffiTaq® Hot Start DNA Polymerase (5 U/μl)
    • AffiTaq® Buffer (10X)
    • MgCl2 (if not included in the buffer, add as specified in the product instructions)
    • Sterile, nuclease-free water

    Note: Exclude dNTPs from the master mix as they will be added separately.

PCR Reaction Setup:

  1. Set up the PCR reaction tubes/strips on ice.
  2. Add the following components to each PCR tube:
Components Concentration Volume per Reaction Final Concentration
AffiTaq® Buffer - dNTP free & Mg2+ free 10X 5 μL 1X
dNTP 10 mM 1 μL 200 μM
AffiTaq® MgCl2 25mM 2 to 8 μL 1 to 4 mM
AffiTaq® Hot Start DNA Polymerase 5U/μL 0.25 to 0.5 μL 1.25 to 2.5 U
Primer Mix 25 2 μL 1X
Template - - Less than 1μg/Reaction
ddH2O - Adjust to 50 μL -
  1. Mix the contents gently by pipetting and briefly spin down the tubes to collect all the liquid at the bottom.

PCR Cycling Conditions:

  1. Start the PCR machine and set the following cycling parameters:

    • Initial Denaturation: 95°C for 2-5 minutes (1 cycle)
    • Denaturation: 95°C for 15-30 seconds
    • Annealing: [Optimal temperature]°C for 15-30 seconds
    • Extension: 72°C for [Extension time based on product guidelines] seconds
    • Final Extension: 72°C for 5-10 minutes (1 cycle)
    • Hold at 4-10°C
  2. Run the PCR according to the specified cycling parameters for the recommended number of cycles for your specific target.

Post-PCR Steps:

  1. Analyze the PCR products using agarose gel electrophoresis or other appropriate methods.
  2. Store the remaining PCR products at -20°C for further analysis or downstream applications.


  • Adhere to aseptic techniques and wear gloves to prevent contamination.
  • Adjust annealing temperature and extension time, if necessary, for specific primer-template combinations.
  • Always refer to the provided AffiTaq® Hot Start DNA Polymerase set instructions for optimal conditions based on your experiment.