- Isolated from a recombinant source
- Supplied with 10X Reaction Buffer
In addition to DNA repair activity, APE 1 is also capable of regulating the DNA binding activity of many transcription factors in vitro by a redox mechanism (Ref-1). As part of this process, APE 1 has been shown to stimulate the DNA binding activity of Fos-Jun heterodimers, Jun-Jun homodimers and Hela cell AP-1 proteins as well as that of several other transcription factors including NF-kB, Myb and members of the ATF/CREB family (7-9).
Product SourceAn E. coli strain which carries the cloned human APE 1 gene.
The following reagents are supplied with this product:
|Store at (°C)||Concentration|
Advantages and Features
- Single cell gel electrophoresis (Comet assay)
- Alkaline elution
- Alkaline unwinding
- Modified nick translation
Properties and Usage
Unit DefinitionOne unit is defined as the amount of enzyme required to cleave 20 pmol of a 34 mer oligonucleotide duplex containing a single AP site* in a total reaction volume of 10 µl in 1 hour at 37°C.
*An AP site is created by treating 20 pmol of a 34 mer oligonucleotide duplex containing a single uracil residue with 1 unit of Uracil-DNA Glycosylase (UDG) for 2 minutes at 37°C.
1X NEBuffer 4
Incubate at 37°C
1X NEBuffer 4:
50 mM Potassium Acetate
20 mM Tris-acetate
10 mM Magnesium Acetate
1 mM DTT
pH 7.9 @ 25°C
10 mM Tris-HCl
50 mM NaCl
1 mM DTT
0.05 mM EDTA
200 μg/ml BSA
pH 8.0 @ 25°C
Heat Inactivation65°C for 20 min
Unit Assay Conditions1X NEBuffer 4 containing 20 pmol of fluorescently labled oligonucleotide duplex in a total reaction volume of 10 μl.
Quality Control AssaysThe 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.
- 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.
- Protein Purity (SDS-PAGE):
The physical purity is assessed by comparing contaminating protein bands in a concentrated sample to the protein of interest band in a sample of known dilution. The purity is determined by SDS-PAGE.
- Recommended Dilution for the Comet Assay: 1:103. For a protocol please visit: http://cometassay.com
- Robson, C.N. and Hickson, D.I. (1991). Nucl. Acids Res.. 19, 5519-5523.
- Demple, B. et al. (1991). Proc. Natl. Acad. Sci. USA. 88, 11450-11454.
- Barzilay, G. et al. (1995). Nucl. Acids Res.. 23, 1544-1550.
- Barzilay, G. et al. (1995). Nature Struc. Biol.. 2, 451-468.
- Gorman, M.A. et al (1997). EMBO J.. 16, 6548-6558.
- Xanthoudakis, S. et al. (1992). EMBO J.. 11, 3323-3335.
- Walker, L.J. et al. (1993). Mol. Cell. Biol.. 13, 5370-5376.
- Vidal, A.E. (2001). EMBO J.. 20, 6530-6539.
- Wilson, D.M. III et al. (1995). J. Biol. Chem.. 270, 16002-16007.
- Flaherty, D.M. (2001). Am. J. Respir. Cell. Mol. Biol.. 25, 664-667.