sho-phire-enzymes

Hot start PCR with improved speed, yields, and amplicons length

Phire™ Hot Start II DNA Polymerase incorporates a dsDNA-binding domain that allows short extension times (10-15 s/kb), helps improve yields, and can increase fidelity two-fold compared to Taq DNA polymerase. In addition, the unique hot start technology allows complete reactivation of the enzyme in “zero-time” at standard cycling temperatures. This combination of features makes Phire Hot Start II DNA Polymerase an ideal solution for routine and high throughput PCR applications.


Features:

  • Very fast enzyme with quick hot start (no reactivation step required)
  • Minimal reaction optimization due to high inhibitor tolerance
  • Higher yields and amplicons length than with any hot start Taq
  • Direct loading on gels with green reaction buffer formats

Complete PCR cycling in less than half the time

Phire Hot Start II DNA Polymerase requires extension time of only 10 s/kb, has zero-time reactivation step and allows direct loading of PCR products on gels. Due to these features PCR protocols with Phire Hot Start II DNA Polymerase can be significantly faster than protocols with Taq DNA polymerases.

 

Short cycling times with Phire Hot Start II DNA Polymerase.  A 600 bp fragment from human genomic DNA was amplified with five different hot start DNA polymerases. With Phire Hot Start II DNA Polymerase, the PCR protocol was completed up to four times faster than with Taq DNA polymerases utilizing chemically modified or antibody-based hot start technologies (suppliers A-D). Green buffer further reduces experimental time by eliminating one pipetting step and allowing for direct loading on gel.

Green reaction buffer for ultimate convenience

Phire Green Hot Start II DNA Poylmerase and PCR Master Mixes are supplied with 5x Phire Green Reaction Buffer that includes a density reagent and two tracking dyes for direct loading of PCR products on gels.

Reaction mixtures containing Phire Green Reaction Buffer. (A) Before electrophoresis. (B) Blue and yellow tracking dyes after electrophoresis.  

Superior yields in significantly shorter time

Phire Hot Start II DNA Polymerase incorporates a dsDNA-binding domain that allows very short cycling times and significantly improves amplicons yields. High yields can be achieved with both Phire Hot Start II stand-alone enzyme and PCR master mix formats.

Superior target yields with Phire Hot Start II DNA Polymerase.  A 1.5 kb fragment from the human cathepsin K gene was amplified with different hot start DNA polymerases. Phire Hot Start II DNA Polymerase amplified high amounts of specific PCR product in just 29 minutes. In contrast, the PCR protocols for hot start Taq DNA polymerases were substantially longer and resulted in lower product yields.


Superior target yields with Phire Hot Start II PCR Master Mix.
A 2 kb fragment of human β-globin gene was amplified with different hot start PCR master mixes. Phire Green Hot Start II PCR Master Mix delivered high yields of specific product in just 41 minutes whereas other master mixes provided lower yields and in most cases required longer protocols.

Higher amplicon length than with any hot start Taq DNA polymerase

Phire Hot Start II DNA Polymerase can amplify amplicons longer than any hot start Taq DNA polymerase. Up to 7.5 kb of gDNA and 20 kb of phage DNA can be amplified with Phire Hot Start II stand-alone enzyme and PCR master mix formats.

Higher amplicons length with Phire Hot Start II DNA Polymerase. Five genomic DNA fragments of different lengths were amplified with three different hot start DNA polymerases. Phire Hot Start II DNA Polymerase produced all five amplicons with very high yields whereas hot start Taq DNA polymerases produced significantly lower yields and failed to amplify the 7.5 kb fragment.
1: 0.6 kb, 2: 1.0 kb, 3: 1.5 kb , 4: 2.7nbsp; kb, 5: 7.5 kb

Higher amplicons length with Phire Hot Start II PCR Master Mix. Five genomic DNA fragments of different lengths were amplified with hot start PCR master mixes. Phire Green Hot Start II PCR Master Mix produced all five amplicons with high yields. Other hot start PCR master mixes produced lower or no yields, with some also amplifying non-specific products.
1: 0.47 kb, 2: 1.1 kb, 3: 1.7 kb, 4: 3.5 kb, 5: 7.5 kb