Antibody‑mediated hot‑start for high specificity
Antibody-based Hot-Start Taq DNA Polymerase ensures exceptional specificity and yield in demanding PCR applications. Inactive during setup and rapidly activated by heat, it minimizes primer-dimer formation and nonspecific amplification, enabling clean, reproducible results across diverse templates up to 5 kb.
| Product Number | EM15HSDNA |
| Function | Antibody-mediated hot start; Release on denaturation |
| For Use In | Room-temp setup, high-specificity PCR |
| Molecular Weight | 94 kDa |
| Concentration | 5 U/µL |
| Delivery Format | Tubes |
| Intended Use | This product is intended for research use only |
| Volume | 50 µL 4 x 250 µL |
| Source | Recombinant E. coli |
| Shipping Conditions | Cold Pack |
| Storage Temperature | -20°C |
| Stability | ≥12 months |
Hot Start DNA Polymerase is a highly pure, thermostable enzyme designed for demanding PCR applications requiring exceptional specificity, sensitivity, and robustness. The enzyme utilizes antibodies to block polymerase activity at ambient temperatures, which dissociate after the initial denaturation step at 95°C. This automatic 'hot start' mechanism enhances sensitivity, specificity, and yield, while also allowing for reaction assembly at room temperature. Optimized for the amplification of 0.1–5 kb DNA targets, this enzyme delivers high yields, even from low copy number templates. Its advanced formulation includes an optimized buffer system and compatibility with high-quality dNTPs, ensuring reliable performance across a broad range of amplicons. Hot Start DNA Polymerase is resistant to PCR inhibitors, such as blood (up to 20%), ethanol, and humic acid, making it ideal for challenging samples with carry-over inhibitors. Additionally, it is well-suited for GC-rich templates (up to 70%). This enzyme is a versatile tool for routine PCR, RT-PCR, and TA cloning, offering excellent specificity and sensitivity for both standard and demanding workflows.
We offer flexible customization to meet your specific needs:
One unit is defined as the amount of enzyme required to catalyze the incorporation of 10 nmol of dNTP into acid-soluble form in 30 minutes at 72°C in the presence of the reaction buffer.
Functional assay
Human genomic DNA was amplified using the DNA Polymerase and specific primers to produce a distinct band of 750 bp.
Self-priming activity
Standard PCR is carried out without primers, using the DNA Polymerase and human genomic DNA. No products were amplified.
Exonuclease assay
Linearized lambda/HindII fragments are incubated with the DNA Polymerase in a 50 µl reaction mixture for 4 h at 37°C. No degradation of DNA was observed.
Endonuclease assay
lambda DNA is incubated with the DNA Polymerase in a 50 µl reaction mixture for 4 h at 37°C. No degradation of DNA was observed.
Nick Activity
Supercoiled plasmid DNA is incubated with the DNA Polymerase in a 50 µl reaction mixture for 4 h at 37°C. No conversion of covalently closed circular DNA to nicked DNA was detected.
Contamination with E. coli DNA
A sample of the denatured DNA Polymerase is analyzed with specific primers targeting the 16S rRNA gene in qPCR for the presence of contaminating E. coli DNA. No E. coli was detectable.
The optimal primer concentration depends on the application, but 0.2–1 µM of each primer (forward and reverse) is commonly used in PCR reactions and is also the range recommended in the user guide. We highly recommend optimizing primer concentration based on your experimental design. Using too little primer can reduce amplification efficiency and yield, while excessive primer concentrations may increase nonspecific amplification and primer-dimer formation.
The enzyme and its buffer system are optimized for a slightly higher melting temperature, providing researchers with a starting point designed to improve specificity and sensitivity for assays in general.
Since each assay is unique, we strongly recommend performing a gradient PCR as part of the optimization to identify the most suitable annealing temperature for your specific design.
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