rE. coli MutY with Buffer

4000-500-EB Datasheet / COA / SDS

Discontinued Product

4000-500-EB has been discontinued.
View all MutY DNA Glycosylase products.
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rE. coli MutY with Buffer Summary

Kit containing rE. Coli MutY and a buffer.

Product Description

E. coli MutY DNA Glycosylase acts together with E. coli Formamidopyrimidine-DNA Glycosylase, or Fpg, to prevent the potentially mutagenic consequences of 8-oxo-dG lesions. 8-oxo-dG lesions escaping repair by Fpg frequently pair with Adenine (A) during DNA replication, producing an 8-oxo-dG:A mispair. MutY removes the A from this base pair to initiate base excision repair. In the absence of MutY, DNA replication after an 8-oxo-dG:A mismatch results in thymine incorporation opposite the adenine in one of the daughter strands, creating a fixed mutation. MutY has an associated AP lyase activity.

Assay Conditions and Analysis

4 pmoles of A/G mismatch oligonucleotide set with the A oligo end-labeled, 1X REC Buffer 4 (10 mM HEPES-KOH (pH 7.4), 100 mM KCl, 10 mM EDTA), and serial dilutions of enzyme in a 20 μL reaction volume are incubated for 1 hour at 37 °C. To complete cleavage of a basic site, fresh NaOH is added to final concentration of 166 mM then heated for 15 minutes at 95 °C. For analysis, 24 μL of 2X Loading Buffer (20 mM EDTA, 97% formamide, and 0.2% bromophenol blue) is added, the samples are heated at 95 °C for 5 min then fast cooled to 2-8 °C, and the cleavage products are resolved by 20% denaturing polyacrylamide gel electrophoresis. The bands are analyzed to quantify the cleavage products.

Kit Contents

• 10X REC™ Buffer 4
• E. coli Mut Y DNA Glycosylase

Specifications

Shipping Conditions
The product is shipped with dry ice or equivalent. Upon receipt, store it immediately at the temperature recommended on the product label.
Storage
Store the unopened product at -70 °C. Use a manual defrost freezer and avoid repeated freeze-thaw cycles. Do not use past expiration date.
Species
E. coli

Limitations

For research use only. Not for diagnositic use.

Product Datasheets

Reagents Provided

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