Description: Exonuclease III (E. coli) catalyzes the stepwise removal of mononucleotides from 3`- hydroxyl termini of double stranded DNA. Exonuclease III activity depends partially on helical structure and displays sequence dependence (C > A = T > G). Temperature, salt concentration and the ratio of enzyme to DNA greatly affect enzyme activity, requiring reaction conditions to be tailored to specific applications.
Applications:
- unidirectional nested deletions;
- site-directed mutagenesis;
- preparation of strand-specific probes;
- preparation of single-stranded substrates for dideoxy sequencing.
Unit
One unit is the amount of enzyme required to produce 1 nmol of acid soluble total nucleotide in 30 minutes at 37oC.
Unit Assay Conditions: 50 mM Tris-HCl (pH 7.5 at 25oC); 1 mM MgCl2; 1 mM DTT, 0.15 mM sonicated pancreatic DNA.
Reaction buffer
SE-buffer Exonuclease III, (50 mM Tris-HCl (pH 7.6 at 25°C); 1 mM MgCl2.)
Optimal temperature
37oC
Storage conditions
10 mM Tris-HCl (pH 7.5); 50 mM KCl; 0.5 mM EDTA; 1 mM 2-mercaptoethanol; 50% glycerol. Store at -20oC.
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