| Source: |
Purified from E. coli containing a recombinant plasmid harboring the E. coli MutY gene. |
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| Unit Definition: |
One unit is the amount of enzyme required to cleave 1 pmole of an oligonucleotide duplex containing an A/G mismatch in 1 hour at 37°C. Only the strand with the A is cleaved.
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| Assay Conditions: |
1X RECTM Buffer 9 (10 mM HEPES-KOH (pH 7.4), 100 mM KCl, 1 mM EDTA, 1 mM EGTA, and 0.1 mM DTT), 4 pmoles 32P hypoxanthine oligonucleotide, 4 pmoles complementary oligonucleotide, and serial dilutions of enzyme in a reaction volume of 20 µl. Incubate for 1 hour at 37°C. Reaction products are resolved by 20% denaturing PAGE.
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| Applications: |
Mismatch cleavage assays
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| Storage: |
Freeze in working aliquots at -20°C in a manual defrost freezer to avoid repeated freeze-thawing. Enzyme is supplied in 10 mM HEPES-KOH (pH 7.4), 100 mM KCl, 1 mM EDTA, 0.1 mg/ml BSA, and 50%(v/v) glycerol. |
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| Specificity: |
MutY DNA glycosylase recognizes A/G and A/8-oxo-dG mismatches in duplex DNA and cleaves the strand containing the A. The opposite strand is not cleaved. |
