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

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:

Product Specifications:

Composition:

• 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.

Applications:

• 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

Protocol:

Preparation:

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:

3. 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.

Notes:

• 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.