Therminator™ DNA Polymerase

Description

Therminator DNA Polymerase is a 9°N™ DNA Polymerase variant with an enhanced ability to incorporate modified substrates such as dideoxynucleotides, ribonucleotides and acyclonucleotides (1,2).

Highlights

  • Isolated from a recombinant source
  • Supplied with 10X Reaction Buffer
  • Enhanced incorporation of modified nucleotides
  • Single base incorporation of dideoxy and acyclonucleotides

Product Source

An E. coli strain that carries the 9°N (D141A / E143A / A485L) DNA Polymerase gene, a genetically engineered form of the native DNA polymerase from Thermococcus species 9°N-7.

Reagents Supplied

The following reagents are supplied with this product:

Store at (°C)Concentration
ThermoPol® Reaction Buffer Pack-2010X

Advantages and Features

Applications

  • DNA sequencing by partial ribosubstitution (3)
  • DNA sequencing using dideoxy (4) or acyclo (5) chain terminators
  • SNP analysis with dideoxy or acyclo chain terminators (6)

Properties and Usage

Unit Definition

One unit is defined as the amount of enzyme that will incorporate 10 nmol of dNTP into acid insoluble material in 30 minutes at 75°C.

Reaction Conditions

1X ThermoPol® Reaction Buffer

1X ThermoPol® Reaction Buffer:
20 mM Tris-HCl
10 mM (NH4)2SO4
10 mM KCl
2 mM MgSO4
0.1% Triton® X-100
pH 8.8 @ 25°C

Storage Temperature

-20°C

Storage Conditions

10 mM Tris-HCl
100 mM KCl
1 mM DTT
0.1 mM EDTA
50% Glycerol
pH 7.4 @ 25°C

Heat Inactivation

No

Molecular Weight

Theoretical: 90000 daltons

5' - 3' Exonuclease

No

3' - 5' Exonuclease

No

Strand Displacement

+

Unit Assay Conditions

1X ThermoPol Reaction Buffer, 200 µM dNTPs including [3H]-dTTP and 15 nM primed single-stranded M13mp18.

References

  1. Gardner, A.F. and Jack, W.E. (1999). Nucl. Acids Res.. 27, 2545-2555.
  2. Gardner, A.F. and Jack, W.E. (2002). Nucl. Acids Res. 30, 605-613.
  3. Barnes, W.F. (1978). J. Mol. Biol.. 119, 83-99.
  4. Sanger, F., Nicklen, S. and Coulson, A.R. (1977). Proc. Natl. Acad. Sci. USA. 74, 5463-5467.
  5. Trainor, G.L. (1996). U.S. Patent No. 5,558,991.
  6. Haff, L.A. and Simirnov, I.P. (1997). Genome Methods. 7, 378-388.

FAQs

  1. What is the specific activity of Therminator DNA Polymerase?
  2. Can Therminator DNA Polymerase be used in other buffers?
  3. When should Therminator DNA Polymerase be used?
  4. Can Therminator DNA Polymerase be substituted for ThermoSequenase?
  5. Can Therminator DNA Polymerase be used for sequencing?
  6. Why are some of the A's in my sequence weak when using Therminator DNA Polymerase?
  7. Does Therminator DNA Polymerase have 3'→5' proofreading exonuclease activity?

Datacards

The Product Summary Sheet, or Data Card, includes details for how to use the product, as well as details of its formulation and quality controls. The following file naming structure is used to name the majority of these document files: [Catalog Number]Datasheet-Lot[Lot Number]. For those product lots not listed below, please contact NEB at info@neb.com or fill out the Technical Support Form for appropriate document.

Selection Charts

Usage Guidelines & Tips

Troubleshooting Guides

Interactive Tools

Quality Control

Quality Control Assays

The following Quality Control Tests are performed on each new lot and meet the specifications designated for the product. Individual lot data can be found on the Product Summary Sheet/Datacard or Manual which can be found in the Supporting Documents section of this page. Further information regarding NEB product quality can be found here.
  • Endonuclease Activity (Nicking):
    The product is tested in a reaction containing a supercoiled DNA substrate. After incubation for 4 hours the percent converted to the nicked form is determined by agarose gel electrophoresis.
  • Exonuclease Activity (Radioactivity Release):
    The product is tested in a reaction containing a radiolabeled mixture of single and double-stranded DNA. After incubation for 4 hours the exonuclease activity is determined by the % release of radioactive nucleotides.

Certificate of Analysis

The Certificate of Analysis (COA) is a signed document that includes the storage temperature, expiration date and quality control's for an individual lot. The following file naming structure is used to name these document files: [Product Number]_[Size]_[Version]_[Lot Number]

Specifications

The Specification sheet is a document that includes the storage temperature, shelf life and the specifications designated for the product. The following file naming structure is used to name these document files: [Product Number]_[Size]_[Version]

Safety Data Sheet

The following is a list of Safety Data Sheet (SDS) that apply to this product to help you use it safely.

Datacards

The Product Summary Sheet, or Data Card, includes details for how to use the product, as well as details of its formulation and quality controls. The following file naming structure is used to name the majority of these document files: [Catalog Number]Datasheet-Lot[Lot Number]. For those product lots not listed below, please contact NEB at info@neb.com or fill out the Technical Support Form for appropriate document.