Tma Endonuclease III

Description

Tma Endonuclease III is a thermostable homolog of E. coli Endonuclease III, the product of the E. coli nth gene (1). It possesses both N-glycosylase and AP-lyase activities. The N-glycosylase activity releases damaged pyrimidines from double-stranded DNA, generating an abasic site. The AP-lysase activity cleaves the phosphodiester bond 3´ to the abasic site resulting in a 3´-α, β-unsaturated aldehyde and a 5´-phosphate.

Damaged bases recognized and removed by Tma Endonuclease III include 5,6 dihydroxythymine, thymine glycol and abasic sites (2).

Product Source

Cloned from Thermotoga maritima and overexpressed in E. coli.

Reagents Supplied

The following reagents are supplied with this product:

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

Advantages and Features

Applications

  • Alkaline elution (3)
  • Alkaline unwinding (4)

Properties and Usage

Unit Definition

One unit is defined as the amount of enzyme required to cleave 1 pmol of a 60-mer oligonucleotide duplex containing a single AP site* in a total reaction volume of 10 µl in 1 hour at 65°C. 

*An AP site is created by treating 10 pmol of a 60-mer oligonucleotide duplex containing a single uracil residue with 1 unit of Uracil-DNA Glycosylase (UDG) for 2 minutes at 37°C. .

Reaction Conditions

1X ThermoPol® Reaction Buffer
Incubate at 65°C

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

Usage Concentration

10,000 units/ml

Storage Temperature

-20°C

Storage Conditions

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

Heat Inactivation

No

Unit Assay Conditions

1X ThermoPol Reaction Buffer containing 5 pmol of fluorescently labeled oligonucleotide duplex in a total reaction volume of 10 µl.

Notes

  1. Diluent Compatibility: Diluent D
    100 mM KCl, 10 mM Tris-HCl (pH 7.4 @ 25°C), 0.1 mM EDTA, 1 mM DTT, 0.1% Triton X-100 and 50% glycerol.

References

  1. Nelson, K.E. et al. (1999). Nature. 399
  2. Marks, K. unpublished observation.
  3. Pflaum, M. et al. (1998). Free Rad. Res. 29
  4. Hartwig, A. et al. (1996). Toxicol. Lett. 88

FAQs

  1. What is the activity of Tma Endonuclease III in the NEBuffers 1-4?
  2. What is the activity of Tma Endonuclease III in ThermoPol buffer?
  3. What is the molecular weight of Tma Endonuclease III?
  4. Is Tma Endonuclease III a tagged protein?
  5. What substrate is used to test the Tma Endonuclease III activity?
  6. What types of DNA damage does Tma Endonuclease III act upon?

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

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.
  • Non-Specific DNase Activity (16 hour, DNA):
    The DNA is tested in a reaction under standard reaction conditions. After incubation for 16 hours there is no detectable degradation of the DNA as 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.