rM. Thermoautotrophicus TDG with Buffer
rM. Thermoautotrophicus TDG with Buffer Summary
TDG is a thermostable thymine DNA glycosylase from Methanobacterium thermoautotrophicum. The optimal temperature for the enzyme is 65°C. The enzyme lacks significant AP lyase or endonuclease activity. TDG works effectively in heteroduplex analysis to detect C to T transitions.
TDG enzyme recognizes T/G mismatches in duplex DNA and cleaves the strand with the T. The opposite strand is not cleaved. The enzyme also recognizes G/G mismatches if at least one nearest neighbor is an A or T and nicks one strand or the other. The enzyme exhibits poor AP lyase activity. One Unit is the amount of enzyme required to cleave 1 pmole of an oligonucleotide duplex containing a T/G mismatch in 1 hour at 65 °C. Only the strand containing the T is cleaved.
Assay Conditions and Analysis
4 pmoles of T/G mismatch oligonucleotide set with the T oligo end-labeled, 1X REC Buffer 4 (10 mM HEPES-KOH (pH 7.4), 100 mM KCl, and 10 mM EDTA), and serial dilutions of enzyme in a 20 μL reaction volume are incubated for 1 hour at 65 °C. To complete cleavage of a basic site, fresh 1N 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, 95% formamide, and 0.13% bromophenol blue) are added, and the samples heated at 95 °C for 10 min then fast cooled to 2-8 °C. The cleavage products are resolved by 20% denaturing polyacrylamide gel electrophoresis, and percent cleavage quantified.
• 10X REC™ Reaction Buffer 4
• Thermostable TDG Enzyme
For research use only. Not for diagnositic use.
Citations for rM. Thermoautotrophicus TDG with Buffer
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Label-free and amplified electrogenerated chemiluminescence biosensing for the detection of thymine DNA glycosylase activity using DNA-functionalized gold nanoparticles triggered hybridization chain reaction
Authors: W Bai, Y Wei, Y Zhang, L Bao, Y Li
Anal. Chim. Acta, 2019;1061(0):101-109. 2019
Controllable Autocatalytic Cleavage-Mediated Fluorescence Recovery for Homogeneous Sensing of Alkyladenine DNA Glycosylase from Human Cancer Cells
Authors: LJ Wang, ML Luo, XY Yang, XF Li, Y Wu, CY Zhang
Theranostics, 2019;9(15):4450-4460. 2019
DNA cytosine and methylcytosine deamination by APOBEC3B: enhancing methylcytosine deamination by engineering APOBEC3B.
Authors: Fu Y, Ito F, Zhang G, Fernandez B, Yang H, Chen X
Biochem J, 0;471(1):25-35. 0
Methylcytosine and normal cytosine deamination by the foreign DNA restriction enzyme APOBEC3A.
Authors: Carpenter M, Li M, Rathore A, Lackey L, Law E, Land A, Leonard B, Shandilya S, Bohn M, Schiffer C, Brown W, Harris R
J Biol Chem, 0;287(41):34801-8. 0
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