Thermo Scientific Exonuclease III (ExoIII) exhibits four catalytic activities. The 3'→5' exodeoxyribonuclease activity of ExoIII is specific for double-stranded DNA. ExoIII degrades dsDNA from blunt ends, 5'-overhangs or nicks, releasing 5'-mononucleotides from the 3'-end of the DNA strand, producing single-stranded DNA fragments. It is not active on DNA 3'-overhangs (at least four bases long and without a 3'-terminal C-residue), single-stranded DNA, or phosphorothioate-linked nucleotides.

ExoIII 3'-phosphatase activity removes the 3'-terminal phosphate group to generate a 3'-OH group. ExoIII RNase H activity can exonucleolytically degrade RNA strands in RNA-DNA hybrids. The ExoIII apurinic/apyrimidine-endonuclease activity cleaves the phosphodiester bond at the apurinic or apyrimidine site, producing a 5'-terminus (abasic deoxyribose 5'-phosphate residue).

Product Advantages

• Active in restriction enzyme buffer

Applications

• Works with S1 nuclease to create unidirectional deletions in DNA fragments
• Generation of single-stranded templates for DNA dideoxy sequencing
• Site-directed mutagenesis
• Cloning of PCR products
• Strand-specific probe preparation

Remarks

The DNA cleavage speed of ExoIII depends on the temperature, salt concentration and molar ratio of DNA to enzyme in the reaction mixture. Optimized reaction conditions need to be retested for each experiment.