FAQ List - ThermaStop™

 

How do I re-suspend ThermaStop™ for use?

 

To prepare 5 Units/µl ThermaStop, add 100 µl 10 mM molecular-grade Tris-Cl, pH 8.3 to 500 units dry reagent (500 µl 10 mM Tris-Cl, pH 8.3 to 2500 units dry reagent), vortex 1-2 minutes, then centrifuge briefly. Allow tube to sit at room temperature for 15 minutes with occasional mixing to ensure reagent is completely dissolved.

 

Do I need to centrifuge the material after re-suspension?

 

Yes, centrifuge briefly to bring any liquid to bottom of the tube.

 

How long can I store the dry material?

 

Dried ThermaStop can be stored for at least a year if kept at room temperature in the vial provided.

 

How long can I store the liquid material?

 

Liquid ThermaStop can be stored for up to six months if kept at 4°C or -20°C.

 

What polymerase enzymes work with ThermaStop?

 

ThermaStop is compatible with most DNA-dependent DNA polymerases, including all native and recombinant Taq DNA polymerases, heat-activated Taq DNA polymerases, and Tfi DNA polymerase.  ThermaStop has been tested with the following commercially available enzymes:

 

Non-hot start polymerase enzymes;

a) Invitrogen Recombinant Taq DNA Polymerase

b) Promega Go Taq® DNA Polymerase

c) Syd Labs Taq DNA Polymerase

d) Hain Life Sciences Taq DNA Polymerase

 

Hot start polymerase enzymes;

a) Platinum™ Taq DNA Polymerase

b) Promega Go Taq® DNA Polymerase

c) Applied Biosystems™  AmpliTaq Gold™ DNA Polymerase

d) LGC KlearTaq™ DNA Polymerase

 

Can I add ThermaStop directly to my Master Mix?

 

Yes. It is not necessary to pre-incubate a DNA polymerase with ThermaStop. ThermaStop should be added to the master mix  before adding the primers and DNA targets.

 

How long can I wait after adding ThermaStop before starting my reaction?

 

Once ThermaStop is added to a PCR master mix the reaction can be started right away.  If desired, a PCR master mix with ThermaStop can be stored for up to 24 hours at room temperature prior to PCR.

 

How many units of ThermaStop do I use with my Taq Polymerase?

 

Add an equal number of units of ThermaStop and Taq DNA polymerase to a PCR master mix. For reactions producing large amounts of non-specific products, add up to 1.5-4 units ThermaStop per unit of Taq DNA polymerase.

 

Can I use ThermaStop in an RT-PCR assay?

 

We recommend ThermaStop-RT for all RT-PCR assays.

 

Can I use ThermaStop in a Multiplex PCR assay?

 

Yes. We also recommend using ThermaStop and ThermaGo, a reagent that enhances the specificity of Taq DNA polymerase by suppressing mis-priming during the annealing and extension steps of PCR.

 

What kind of improvement should I see with ThermaStop?

 

ThermaStop will reduce the production of primer dimers and non-specific products and generate higher yields of the correct products (e.g. stronger probe signals, correct bands on gels) compared to non-hot start conditions.

 

 

 

 

 

 

FAQ List - ThermaGo™

 

How do I re-suspend ThermaGo for use?

 

To prepare 5 Units/µl ThermaStop, add 100 µl 10 mM molecular-grade Tris-Cl, pH 8.3 to 500 units dry reagent (500 µl 10 mM Tris-Cl, pH 8.3 to 2500 units dry reagent), vortex 1-2 minutes, then centrifuge briefly. Allow tube to sit at room temperature for 15 minutes with occasional mixing to ensure reagent is completely dissolved.

 

Do I need to centrifuge the material after re-suspension?

 

Yes, centrifuge briefly to bring any liquid to bottom of the tube.

 

How long can I store the dry material?

 

Dried ThermaGo can be stored for at least a year if kept at room temperature in the vial provided.

 

How long can I store the liquid material?

 

Liquid ThermaStop can be stored for up to six months if kept at 4°C.

 

What polymerase enzymes work with ThermaGo?

 

ThermaGo is compatible with all Taq DNA polymerases.

 

Can I add ThermaGo directly to my Master Mix?

 

Yes. It is not necessary to incubate a PCR DNA polymerase with ThermaGo.

 

How long can I wait after adding ThermaGo before starting my reaction?

 

Once ThermaGo is added to a PCR master mix the reaction can be started right away.

 

How many units of ThermaGo do I use with my Taq Polymerase?

 

To use, begin by adding an equal number of units of ThermaGo and Taq DNA polymerase to a PCR master mix. To further optimize, test ThermaGo in the range of 0.5 to ≥2.0 Units per Unit of Taq DNA polymerase. Note: very high levels of ThermaGo can inhibit amplification.

 

Can I use ThermaGo in a Multiplex PCR assay?

 

Yes. We also recommend using ThermaGo together with ThermaStop, a reversible hot start reagent compatible with all Taq DNA polymerases. ThermaStop acts directly on the polymerase to prevent non-specific enzymatic activity below 50°C. Polymerase activity is fully restored at 60°C, but is inhibited again by ThermaStop by cooling the reaction.

 

What kind of improvement should I see with ThermaGo?

 

Reactions containing optimal levels of ThermaGo will exhibit greater quantitative reproducibility due to suppression of non-specific products. Use of ThermaGo will produce higher yields of the intended products (e.g. stronger probe signals, correct bands on gels).

 

THE FUTURE OF PCR

©2017 THERMAGENIX, INC.

THE FUTURE OF PCR