Thermo Scientific™

Phusion Hot Start II High-Fidelity PCR Master Mix

Catalog number:  F565S

Related applications:


Back to top


Thermo Scientific Phusion High-Fidelity DNA Polymerases set a gold standard for high performance PCR. Featuring an error rate 50-fold lower than that of Taq, and 6-fold lower than that of Pfu, Phusion High-Fidelity DNA Polymerase is excellent choice for cloning and other applications requiring high fidelity. Phusion DNA Polymerases offer robust performance with short protocol times, even in the presence of PCR inhibitors, and generate higher yields with lower enzyme amounts than other DNA polymerase.


• Reaction set up at room temperature
• No non-specific amplification and primer degradation during reaction set up
• Zero-time reactivation due to unique hot start technology
• High fidelity (52X Taq)
• Fast PCR due to short extension times (15-30 s/kb)
• Robust reactions, minimal optimization needed
• Increased product yields with minimal enzyme amounts
• Available as a Green buffer format permitting direct loading on gels (F-566S or F-566L)

Using Phusion Hot Start II High-Fidelity DNA Polymerase, amplification proceeds without the production of nonspecific products due to the combination of Phusion DNA Polymerase and a reversibly bound, specific Affibody ligand that inhibits DNA polymerase activity at room temperature. The Affibody ligand also inhibits the 3´→5´ exonuclease activity of the polymerase, thus preventing degradation of primers and template DNA during reaction set up. At temperatures that promote polymerase activity, the ligand is released, rendering the polymerase fully active. Phusion Hot Start II DNA Polymerase is immediately reactivated at high temperatures so it does not require a separate activation step in PCR protocols. Phusion Hot Start II High-Fidelity PCR Master Mix is convenient 2X mix designed to minimize the number of pipetting steps. The master mix contains Phusion Hot Start II DNA Polymerase, nucleotides and optimized reaction buffer including MgCl2. Only template and primers need to be added to the PCR reaction.


• High-fidelity PCR
• High throughput
• Amplification of difficult (GC-rich) templates
• Template generation for sequencing
• Multiplex PCR
• Long-range PCR
• Cloning
• Mutagenesis
• Microarray

Using Phusion DNA Polymerases
Annealing rules for Phusion DNA Polymerases are different from many common DNA polymerases (such as Taq DNA polymerases). For optimal results, use our Tm calculator at

For Research Use Only. Not for use in diagnostic procedures.


Hot Start: Built-In Hot Start
Includes: 2X Phusion Hot Start II High-Fidelity PCR Master Mix (provides 1.5 mM MgCl2 in the final 1X concentration); Water, nuclease-free; DMSO
Quantity: 100 x 50μL rxns
Concentration: 2X
Fidelity (vs. Taq): 52 X
For Use With (Application): PCR Amplification
GC-Rich PCR Performance: Medium
Overhang: Blunt
Polymerase: Phusion Hot Start II High-Fidelity DNA Polymerase
Product Line: Phusion
Product Type Specs: Hot Start High-Fidelity PCR Master Mix
Reaction Format: SuperMix or Master Mix
Reaction Speed: Fast

Contents & storage


• 2X Phusion Hot Start II High-Fidelity PCR Master Mix (provides 1.5 mM MgCl2 in the final 1X concentration)
• Water, nuclease-free

Store at -20°C.