9°Nm™ DNA Polymerase

Description

9°Nm™ DNA Polymerase is a thermophilic DNA polymerase that has been genetically engineered to have a decreased 3'→ 5' proofreading exonuclease activity (1-5% of the wildtype). 9°Nm DNA Polymerase features a half-life of 6.7 hours at 95°C.

9°Nm DNA Polymerase crystals (Ira Schildkraut and Rebecca Kucera, New England Biolabs, Inc.)

Highlights

  • Isolated from a recombinant source
  • Supplied with 10X Reaction Buffer

Product Source

An E. coli strain that carries the 9°N (E143D) DNA Polymerase gene, (1,2) a genetically engineered form of the native DNA polymerase from the extremely thermophilic marine archaea Thermococcus species 9°N-7. The archaea was isolated from a submarine thermal vent, at a depth of 2,500 meters, 9° north of the equator at the East Pacific Rise (3).

Reagents Supplied

The following reagents are supplied with this product:

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

Advantages and Features

Applications

  • Primer extension
  • SNP analysis

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

Yes

Strand Displacement

+

Resulting Ends

  • Mix of blunt and Single-base 3´ Overhangs

Unit Assay Conditions

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

Notes

  1. It is suggested that the number of units be optimized with each primer:template.
  2. MgSO4 is usually optimal at 2 mM, although occasionally a 1 mM level is better. For such reactions, we offer a ThermoPol II (Mg-free) Reaction Buffer Pack (#B9005) to which 1 mM MgSO4 can be added.
  3. Diluent E (#B8005) is available for making dilutions of 9°Nm DNA Polymerase.
  4. Additional buffer packs are also available (#B9004). Each buffer pack contains 4 vials of 10X ThermoPol Reaction Buffer (1.5 ml each) and 1 vial of 100 mM MgSO4.

References

  1. Southworth, M.W. et al. (1996). Proc. Natl. Acad. Sci. USA. 93, 5281-5285.
  2. Rodriquez, A.C. et al. (2000). J. Mol. Biol.. 299, 447-462.
  3. Dr. Holger Jannasch (1991). Thermococcus sp. (strain 9°N-7) isolated. Woods Hole Oceanographic Institute.

FAQs

  1. I can't get 9°Nm DNA Polymerase to work, yet Taq DNA Polymerase works fine.
  2. Are the DNA fragments produced by 9°Nm DNA Polymerase blunt-ended or do they have the single-base 3' overhang that Taq DNA Polymerase yields?
  3. Can I use 9°Nm DNA Polymerase to polish the ends of DNA fragments?
  4. Can 9°Nm DNA Polymerase be used in other buffers?

Tech Tips

For reactions requiring Mg2+ optimization, ThermoPol II (Mg-Free) Reaction Buffer Packs are available (NEB# B9005S).

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.

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.