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| Product Name | Gold High-Fidelity DNA Polymerase |
| Catalog No. | IVDRM-0103 |
| Description | This enzyme is a genetically engineered high-fidelity DNA polymerase derived from Pyrococcus furiosus DNA polymerase. It possesses 5'→3' DNA polymerase activity and 3'→5' exonuclease (proofreading) activity, achieving a fidelity 52-fold higher than Taq DNA polymerase and 6-fold higher than conventional Pfu DNA polymerase. A hot-start factor in the enzyme formulation markedly improves amplification detection rates and product specificity, while an added elongation factor provides long-fragment capability with extension rates of up to 15 sec/kb and target lengths of up to 10 kb. The optimized buffer contains PCR-enhancing components, making the enzyme well suited to complex templates. Amplification products are blunt-ended. |
| Intended Use | High-fidelity PCR amplification for gene cloning, site-directed mutagenesis, and other applications requiring accurate amplification of fragments of different lengths and GC contents, including complex templates. |
| Principle / Technology | Engineered from Pyrococcus furiosus (Pfu) DNA polymerase; fidelity arises from 3'→5' exonuclease proofreading activity that corrects misincorporated bases, combined with 5'→3' polymerase activity. A hot-start factor enhances specificity, and an elongation factor supports fast, long-fragment amplification. |
| Sample Type | Genomic DNA (e.g., human, mouse, rice, Arabidopsis), cDNA, plasmid DNA, and λ DNA templates; suitable for fragments of varying lengths and GC contents (30%–70%). |
| Performance Range / Specifications | Fidelity: 52× Taq DNA polymerase and 6× conventional Pfu DNA polymerase; error rate approximately 0.00039% Extension rate: up to 15 sec/kb; extreme speed of 1 sec/kb Amplification length: targets up to 10 kb; human gDNA/cDNA ≤10 kb; λ DNA ≤20 kb; verified range up to 20 kb for simple templates and 10 kb for complex templates Sensitivity: amplifies templates as low as 1 pg Annealing temperature: 68°C GC range: 30%–70% Product ends: blunt ends |
| Specificity | Produces a single, specific target band with higher specificity and yield than comparable high-fidelity polymerases. |
| Reaction Conditions / Protocol | Recommended annealing temperature: 68°C. Extension rate: up to 15 sec/kb (extreme speed 1 sec/kb); effective amplification demonstrated across extension speeds of 1–60 sec/kb. |
| Components / Formulation | Gold High-Fidelity DNA Polymerase (2 U/μL) 2× PCR Buffer (contains Mg2+ and dNTPs) |
| Storage Conditions | Store at -20°C. |
| Shelf Life | 2 years. |
| Package Specifications | 100 U 500 U 1000 U 40000 U |
| Product Form | Standalone enzyme |
| Concentration | 2 U/μL |
| Source / Origin | Genetically engineered enzyme derived from Pyrococcus furiosus (Pfu) DNA polymerase. |
| Shipping Conditions | Shipped with ice packs. |
| Expiration Date / Stability | Valid for 2 years when stored at -20°C. |
| Compatibility | Efficiently amplifies fragments of different lengths (1–12 kb demonstrated) and GC contents (30%–70%) from diverse templates, including gDNA, cDNA, plasmid DNA, and λ DNA. |
| Recommended Buffer System | Supplied optimized 2× PCR buffer containing Mg2+, dNTPs, and PCR-enhancing components. |
| Application Notes / Precautions | Fidelity is based on 3'→5' exonuclease proofreading activity; error rate is approximately 0.00039% (Taq: ~0.0002, i.e., about one mutation per 5 kb). Higher fidelity reduces but cannot completely eliminate mutations. The enzyme generates blunt-ended products. To add A-tails for TA cloning: purify the blunt-ended PCR product (≥20 ng/μL), then combine 14.5 μL of product with 2 μL Taq buffer, 3 μL dNTPs, and 0.5 μL Taq polymerase; incubate at 72°C for 10–20 min and purify. Targets within the verified range (up to 20 kb for simple templates, 10 kb for complex templates) are theoretically amplifiable; actual results depend on template and primer quality. If mutations appear in products, check whether the original template carries mutations or has degraded from repeated freeze-thaw cycles or prolonged UV exposure; minimize freeze-thawing and UV exposure of templates. |
For research use only, not for clinical use.
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