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| Download: | | MSDS PDF | |
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- Isolated from a recombinant source
- Supplied with 10X Reaction Buffer
- Robust amplification and product yield
9°N™m 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.
Source:
An E. coli strain that carries the 9°N (E143D) DNA Polymerase gene, (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 (1).
Applications:
- Primer extension
- SNP analysis
ThermoPol Reaction Buffer Pack (10X)
Enzyme Properties
3´ to 5´ Exonuclease: Yes
5´ to 3´ Exonuclease: No
Strand Displacement: Yes
Heat Inactivation:
No
Molecular Weight:
Theoretical: 90 kDa
Specific Activity:
35,000 units/mg
Reaction & Storage Conditions
Reaction Conditions:
1X ThermoPol Reaction Buffer Pack
1X ThermoPol Reaction Buffer Pack:
20 mM Tris-HCl
10 mM (NH4)2SO4
10 mM KCl
2 mM MgSO4
0.1 % Triton X-100
pH 8.8 @ 25°C
Unit Definition:
One unit is defined as the amount of enzyme that will incorporate 10 nmol of dNTP into acid-insoluble material in a total reaction volume of 50 μl in 30 minutes at 75°C in 1X ThermoPol Reaction Buffer with 200��M dNTPs including [3H]-dTTP and 15 nM primed single-stranded M13mp18.
Concentration:
2,000 units/ml
Storage Conditions:
10 mM Tris-HCl
100 mM KCl
1 mM Dithiothreitol
0.1 mM EDTA
50% Glycerol
pH 7.4 @ 25°C
Storage Temperature:
-20°C
Notes
General notes:
- Diluent buffer (NEB #B8005S) is available for making dilutions of 9°Nm DNA Polymerase.
- Additional buffer packs for this product are also available (NEB #B9004S). Each buffer pack contains 4 vials of 10X ThermoPol Reaction Buffer (1.5 ml each) and 1 vial of 100 mM MgS04.
- It is suggested that the number of units be optimized with each primer:template.
- MgS04 is usually optimal at 2 mM, although occasionally a 1 mM level is better. For such reactions, we offer an ThermoPol II (Mg-free) Reaction Buffer Pack (NEB #B9005S) to which 1 mM MgS04 can be added.
FAQs
- I can't get 9°Nm™ DNA Polymerase to work, yet Taq DNA Polymerase works fine.
- 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?
- Can I use 9°Nm™ DNA Polymerase to polish the ends of DNA fragments?
- Can 9°Nm™ DNA Polymerase be used in other buffers?
Quality Control for Current Lot
Quality control values for a specific lot can be found on the datacard which accompanies each product.
Quality Assurance Statement:
Purified free of contaminating endonucleases and exonucleases.
Endonuclease Assay:
Incubation of a 50 μl reaction in ThemoPol Reaction Buffer supplemented with 400 μM each dNTP containing a minimum of 20 units of 9° North modified DNA Polymerase with 1 μg of supercoiled ΦX174 DNA for 4 hours at either 37°C or 75°C results in < 10% conversion to the nicked form as determined by agarose gel electrophoresis.
9°Nm DNA Polymerase crystals (Ira Schildkraut and Rebecca Kucera, New�England Biolabs, Inc.)
References
- Dr. Holger Jannasch, Woods Hole Oceanographic Institute, 1991, unpublished observations.
- Southworth, M.W. et al. (1996) Proc. Natl. Acad. Sci. USA, 93, 5281-5285.
- Rodriquez, A.C. et al. (2000) J. Mol. Biol., 299, 447-462.
Reagents Sold Separately
ThermoPol Reaction Buffer Pack
Companion Products
Deoxynucleotide Solution Mix
Deoxynucleotide Solution Set
Diluent E
Magnesium Sulfate (MgSO4) Solution
ThermoPol DF (Detergent-free) Reaction Buffer Pack
ThermoPol II (Mg-free) Reaction Buffer Pack
ThermoPol Reaction Buffer Pack
Legal
Patents:
New England Biolabs, Inc.: U.S. Patent No. 5,756,334
New England Biolabs, Inc.: U.S. Patent No. 5,352,778