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Home > Products > DNA Modifying Enzymes and Cloning > DNA Repair Proteins > Endonuclease VIII Endonuclease VIII Catalog # Size Concentration Price Qty   M0299L 2,500 units 10,000 units/ml $244.00 M0299S 500 units 10,000 units/ml $61.00 Prices are in US dollars and valid only for US orders. Download: MSDS PDF Isolated from a recombinant source Supplied with 10X Reaction Buffer Description: Endonuclease VIII from E. coli acts as both an N-glycosylase and an AP-lyase. The N-glycosylase activity releases damaged pyrimidines from double-stranded DNA, generating an apurinic (AP site). The AP-lyase activity cleaves 3´ and 5´ to the AP site leaving a 5´ phosphate and a 3´ phosphate. Damaged bases recognized and removed by Endonuclease VIII include urea, 5, 6- dihydroxythymine, thymine glycol, 5-hydroxy-5- methylhydanton, uracil glycol, 6-hydroxy-5, 6-dihydrothymine and methyltartronylurea (1,2). While Endonuclease VIII is similar to Endonuclease III, Endonuclease VIII has β and δ lyase activity while Endonuclease III has β lyase activity. Source: An E. coli strain which carries the cloned nei gene. Applications: Single cell gel electrophoresis (Comet assay) (3,4,5) Alkaline elution (6) Alkaline unwinding (7) Reagents Supplied: Endonuclease VIII Reaction Buffer (10X) Enzyme Properties Nuclease Properties Comparison Heat Inactivation: 75°C for 10 minutes Reaction & Storage Conditions Reaction Conditions: 1X Endonuclease VIII Reaction Buffer Incubate at 37°C. 1X Endonuclease VIII Reaction Buffer: 10 mM Tris-HCl 75 mM NaCl 1 mM EDTA pH 8.0 @ 25°C Unit Definition: One unit is defined as the amount of enzyme required to cleave 1 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 in 1X Endonuclease VIII Reaction Buffer containing 10 pmol of fluorescently labeled oligonucleotide duplex. * An AP site is created by treating 10 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. Concentration: 10,000 units/ml Storage Conditions: 10 mM Tris-HCl 250 mM NaCl 0.1 mM EDTA 50% Glycerol pH 8.0 @ 25°C Storage Temperature: -20°C Notes Usage notes: Recommended Dilution for Comet Assay: 1:104 to 1:105 (3,4,5,8). 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 exonucleases and endonucleases. 16-Hour Incubation: A 50 μl reaction containing 1 μg of λDNA (HindIII digest) and 50 units of Endonuclease VIII incubated for 16 hours at 37ºC resulted in a DNA pattern free of detectable nuclease degradation as determined by agarose gel electrophoresis. Exonuclease Activity: Incubation of a 50 μl reaction containing 10 units of Endonuclease VIII with 1 μg of a mixture of single and double-stranded [3H] E. coli DNA (205 cpm/μg) for 4 hours at 37ºC released < 0.4% of the total radioactivity. References Dizdaroglu, M., Laval, J. and Boiteux, S. (1993) Biochemistry, 32, 12105-12111. Hatahet, Z. et al. (1994) J. Biol. Chem., 269, 18814-18820. Singh, N. et al. (1988) Exp. Cell Res., 175, 184-191. Collins, A. et al. (1993) Carcinogenesis, 14, 1733-1735. Collins, A. et al. (1996) Environ. Health Perspect., 104, 465-469. Pflaum, M. et al. (1998) Free Rad. Res., 29, 585-594. Hartwig, A. et al. (1996) Toxicol. Lett., 88, 85-90. Marks, K., unpublished observations.