PfuUltra™ II Fusion HS DNA Polymerase, Cat# 600670

  • Highest accuracy of any PCR enzyme
  • Includes the ArchaeMaxx® polymerase enhancing factor
  • Great PCR success and reliability
  • High specificity hotstart formulation
  • Extended length capability up to 19 kb
  • 70%-80% quicker time-to-results
  • Ideal for PCR cloning, RT-PCR and site-directed mutagenesis

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 Manual Pfu Ultra II Fusion HS DNA Polymerase
 
 

Our second generation Pfu-based fusion DNA polymerases set a new standard in high-fidelity PCR performance. In our new PfuUltra II fusion HS DNA polymerase, we coupled the polymerase fusion technology with our engineered PfuUltra DNA polymerase, hotstart antibody, and proprietary ArchaeMaxx® PCR enhancing factor to achieve extreme accuracy, excellent reliability, high specificity, and long target-length capability while dramatically reducing overall PCR extension times.


Industry Leading Fidelity

Our PfuUltra II fusion HS DNA polymerase is the new industry standard for ultra-high fidelity. Our fidelity testing method(1) demonstrates that PfuUltra II fusion HS DNA polymerase exhibits an accuracy that is over 3-fold higher than any other proofreading enzyme, and has fidelity equal to that of our original PfuUltra™ Hotstart DNA Polymerase. The calculated PCR error rate for the PfuUltra II enzyme is 1 mistake in 2,500,000 nucleotides which is 20-fold fewer mistakes than with Taq DNA polymerase. Figure 1 shows the high accuracy of the PfuUltra II enzyme as well as that of other proofreading DNA polymerases using our testing method. We continually evaluate DNA polymerase accuracy claims using this published method so that relative differences can be compared.
Figure 1 PfuUltra™ II Fusion HS DNA Polymerase, the Highest Fidelity PCR Enzyme The PfuUltra™ II Fusion HS DNA Polymerase exhibits accuracy 3-fold higher than the Phusion™/iProof™ DNA polymerases and 20-fold higher than Taq DNA Polymerase. The fidelity of each listed enzyme was measured using our validated and referenced fidelity assay (1) (Accuracy is equal to 1/error rate).


Enhanced Processivity

We have dramatically increased the processivity of Pfu-based enzymes by fusing our PfuUltra™ DNA polymerase with a high affinity double-stranded DNA binding domain. This domain serves to better anchor the DNA polymerase, preventing early dissociation from the DNA template. The processivity of our PfuUltra II fusion HS DNA polymerase has improved 12 fold over cloned Pfu polymerase (table 1). When compared in manufacturer’s recommended PCR buffers, the processivity of the PfuUltra II enzyme is more than 5 fold higher that of the other fusion enzymes including Phusion™ and PfxUltima™ eznymes. The improved processivity of fusion enzymes leads to higher PCR yields and faster overall run times, hence saving you time and improving template integrity by minimizing exposure to extreme cycling temperatures.
Table 1. Enhanced Processivity of PfuUltra™ II Fusion HS DNA polymerase


Shorter Run Time

The improved speed means 70-80% quicker time-to-results and increased throughput. While typical proofreading DNA polymerases require 1-2 minutes per kilobase extension times, the PfuUltra Il enzyme allows the use of extension time as short as 15 sec/kb. The use of shorter extension times means that PCR cycling takes only one-third to one-fifth of the time required for non-fusion DNA polymerases. As shown figure 2, you can save 1 ½, 5 ½, and 11 hours when amplifying target sizes of 2.6 kb, 6 kb, and 12 kb, respectively. Now you are limited only by the ramping and heating capability of the thermal cyclers.
Figure 2 PfuUltra™ II Enzyme demonstrates unrivaled speed, leading to dramatically shorter overall reaction times. The reaction protocol for PfuUltra™ II enzyme was compared to another proofreader and Taq DNA polymerases. Each protocol was designed to amplify 2.6, 6, and 12 kb products in 30 cycles of 3-step with the recommended extension times for each enzyme.


The ArchaeMaxx® Factor Advantage

In addition to higher accuracy, PfuUltra II fusion HS DNA polymerase provides superior overall performance and more robust amplification of longer targets due to Stratagene's patented ArchaeMaxx® polymerase-enhancing factor. The ArchaeMaxx factor improves the yield of products that contain Pfu DNA polymerase by overcoming "PCR poisoning", which is caused by dUTP accumulation during PCR through dCTP deamination(1). Incorporation of dUTP inhibits Pfu and many other archaeal DNA proofreading polymerases, limiting their efficiency(2). The ArchaeMaxx factor functions as a dUTPase, converting poisonous dUTP to harmless dUMP and inorganic pyrophosphate resulting in greatly enhanced overall PCR performance, including shorter extension times, higher yield and greater target length capability.


Extreme Reliability

The improved processitivity of our PfuUltra II fusion HS DNA polymerase, together with our proprietary ArchaeMaxx factor, enhance the reliability of its performance. Our exclusive ArchaeMaxx factor eliminates dUTP poisioning which causes all other proofreading enzymes to stall(3). Our PfuUltra II polymerase is ideal for amplifying diverse clone collections which demand high fidelity, specificity, and throughput (figure 3).


Amplify Long Targets in a Fraction of the Time

The PfuUltra II fusion HS DNA polymerase amplifies the widest variety of template sizes compared to other proofreading DNA polymerases. Because it requires short cycling times, genomic templates up to 19 kb in length can be amplified in a fraction of the time (Figure 4). For example, the 19 kb target shown in figure 4 can be amplified in 5 hrs, as compared to > 19 hrs required for traditional proofreading enzymes. Compared to the PfuUltra II enzymes, most other commercial fusion enzymes exhibit limited target-length capability, such as up to 6 kb as observed for Phusion DNA polymerase (figure 4). It is critical for you to use high fidelity enzymes for amplifying long targets, since mutation frequency increases linearly with amplicon size.


Improved Specificity with Hot-Start Formulation

PfuUltra II fusion HS DNA polymerase improves specificity, reduces background and enhances yield of many challenging PCR systems. This unique hot start formulation incorporates monoclonal antibodies that effectively neutralize both the DNA polymerase and 3’-5’ exonuclease activities until cycling has begun. PfuUltra II fusion HS DNA polymerase is ideal when amplifying low-copy-number targets from complex genomic DNA or when PCR reactions sit at room temperature for prolonged periods of time.