Exonuclease III.
Exonuclease III (EC catalyzes the stepwise removal of mononucleotides from 3´-OH termini of dsDNA. The preferred substrates are blunt or recessed 3´ termini, although the enzyme also acts at nicks in dsDNA to produce ss gaps. The enzyme is not active on ssDNA, and thus 3´-protruding termini (longer as 4 nt) are resistant to cleavage. This property used to produce unidirectional deletions from a linear molecule with one resistant (3´-overhang) and one susceptible (blunt or 5´-overhang) terminus. Unidirectional deletions can also be created if one terminus contains a-phosphorothioated nucleotide.
Unit definition

1 unit produces 1 nmol of acid soluble nucleotides from dsDNA in 30 min at 37°C.

Reaction buffer
#B27 (10x): 660 mM Tris-HCl pH8.0, 6,6 mM MgCl2.
Reaction buffers
Digestion rate depends on reaction temperature, sequence, ionic strength and enzyme-to-DNA ratios. Mononucleotides are released as follow: C>A,T>G; digestion rate varies from 80bp/min at 25°C to 420 bp/min at 37°C.
Store at -20°C.
unidirectional nested deletions, site-directed mutagenesis, preparation of strand-specific probes.

Quality control

activity, absence of contaminating endo- and exonucleases, specific performance tests.


1. Rogers, S.G., Weiss, B. (1980) Meth. Enzymology 65, 201. 2. Henikoff, S. (1984) Gene 28, 351-359. 3. Guo, L.H, Wu, R. (1982) Nucl. Acids Res. 10, 2065-2084. 4. Vandeyar, M.A. (1988) Gene 65, 129-133.

Catalog #





Pack size


20.000 U





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