To access your account, log in or register.	shopping cart | log in

FAQs for Exonuclease III (E. coli) FAQs for DNA Modifying Enzymes and Cloning Technical Reference for DNA Modifying Enzymes and Cloning Enzyme Finder NEBcutter NEBuffer Chart Double Digest Finder Isoschizomers DNA Sequences and Maps REBASE NEBuffer 1

Home > Products > DNA Modifying Enzymes and Cloning > Nucleases > Exonuclease III (E. coli) Exonuclease III (E. coli) Catalog # Size Concentration Price Qty   M0206L 25,000 units 100,000 units/ml $212.00 M0206S 5,000 units 100,000 units/ml $53.00 Prices are in US dollars and valid only for US orders. Download: MSDS PDF Isolated from a recombinant source 3'→5' exonuclease Produces unidirectional nested deletions Site-directed mutagenesis Supplied with 10X Reaction Buffer Description: Catalyzes the stepwise removal of mononucleotides from 3´-hydroxyl termini of duplex DNA (1). A limited number of nucleotides are removed during each binding event, resulting in coordinated progressive deletions within the population of DNA molecules (2). The preferred substrates are blunt or recessed 3´-termini, although the enzyme also acts at nicks in duplex DNA to produce single-strand gaps. The enzyme is not active on single-stranded DNA, and thus 3´-protruding termini are resistant to cleavage. The degree of resistance depends on the length of the extension, with extensions 4 bases or longer being essentially resistant to cleavage. This property can be exploited to produce unidirectional deletions from a linear molecule with one resistant (3´-overhang) and one susceptible (blunt or 5´-overhang) terminus (3). Exonuclease III activity depends partially on helical structure (4) and displays sequence dependence (C>A=T>G; ref. 5). Temperature, salt concentration and the ratio of enzyme to DNA greatly affect enzyme activity, requiring reaction conditions to be tailored to specific applications.  Exonuclease III has also been reported to have RNase H, 3´-phosphatase and AP-endonuclease activities (1). Source: Purified from E.coli K-12, BE257/pSGR3 strain (kindly supplied by B. Weiss) Applications: Unidirectional nested deletions (3) Site-directed mutagenesis (6) Preparation of strand-specific probes (2) Preparation of single-stranded substrates for dideoxy sequencing (7) Reagents Supplied: NEBuffer 1 (10X) Enzyme Properties Heat Inactivation: 70°C for 20 minutes Specific Activity: 150,000 units/mg Reaction & Storage Conditions Reaction Conditions: 1X NEBuffer 1 Incubate at 37°C. 1X NEBuffer 1: 10 mM Bis-Tris-Propane-HCl 10 mM MgCl2 1 mM Dithiothreitol pH 7.0 @ 25°C Unit Definition: One unit is defined as the amount of enzyme required to produce 1 nmol of acid-soluble total nucleotide in a total reaction volume of 50 μl in 30 minutes at 37°C in 1X NEBuffer 1 with 0.15 mM sonicated duplex [3H]-DNA. Concentration: 100,000 units/ml Storage Conditions: 5 mM KPO4 200 mM KCl 5 mM 2-Mercaptoethanol 0.05 mM EDTA 200 µg/ml BSA 50% Glycerol pH 6.5 @ 25°C Storage Temperature: -20°C Notes Usage notes: Phosphorothioate linkages are not cleaved by Exonuclease III. Unidirectional deletions can also be created by protecting one terminus by incorporation of α-phosphorothioate-containing nucleotide (8). FAQs What is the activity of Exonuclease III in the NEBuffers? Can DNA be blunted using Exonuclease III? Why didn't the reaction using Exonuclease III work? Why does all of the DNA get degraded in my Exonuclease III reaction? How do T7 Exonuclease (NEB# M0263)and Lambda Exonuclease (NEB# M0262) differ from Exonuclease III? Can Exonuclease III be heat inactivated? What is the specific activity of Exonuclease III? Quality Control for Current Lot Quality control values for a specific lot can be found on the datacard which accompanies each product. Endonuclease Activity: Incubation of a 50 μl reaction containing 250 units of Exonuclease III (E. coli) with 1 μg of ΦX174 RF I DNA for 4 hours at 37ºC resulted in < 50% conversion to RFII as determined by agarose gel electrophoresis. References Rogers, G.S. and Weiss, B. (1980) L. Grossman and K. Moldave (Eds.), Methods Enzymol., 65, pp. 201-211. New York: Academic Press. Sambrook, J., Fritsch, E.F. and Maniatis, T. (1989) Molecular Cloning: A Laboratory Manual (2nd. Ed.), 5.84-5.85. Henikoff, S. (1984) Gene, 28, 351-359. Richardson, C.C. et al. (1964) J. Biol. Chem., 239, 251-258. Linxweiler, W. and Horz, W. (1982) Nucl. Acids Res., 10, 4845-4859. Vandeyar, M.A. (1988) Gene, 65, 129-133. Guo, L.H. and Wu, R. (1982) Nucl. Acids Res., 10, 2065-2084. Putney, S., Benkovic, S. and Schimmel, P. (1981) Proc. Natl. Acad. Sci. USA, 78, 7350-7354. Reagents Sold Separately NEBuffer 1