About GpC Methylase (M.CviPI)
Zymo Research's GpC Methylase completely methylates all cytosines (C5) within a 5'…GpC…3'. context in double-stranded DNA. The enzyme is specific for both non-methylated and hemimethylated DNA. The recombinant GpC Methylase is isolated from an E. Coli strain that expresses the methyltransferase gene from Chlorella virus. The reaction conditions are optimized to maximize the processivity of the enzyme to ensure rapid, complete, and reproducible methylation of DNA for accurate DNA methylation analysis. This product is supplied with 10X Reaction Buffer and S-adenosylmethionine cofactor.
|Format||Provided in solution (4 units/µl) w/ 10X Reaction Buffer and 20X SAM|
|Source||Recombinant GpC Methylase is isolated from E. Coli expressing the methyltransferase gene from a Chlorella virus.|
|Storage||Store at -20°C for up to 12 months. Avoid repeated freeze/thawing of reagents. Prolonged storage is at ≤ -70°C.|
|Enzyme Commission Number||(EC 22.214.171.124)|
|Unit Definition||One unit is defined as the amount of enzyme required to "protect" 1 µg of λ DNA against cleavage by HaeIII restriction endonuclease in a total reaction volume of 20 µl for 1 hour at 37°C.|
|Reaction Conditions||GpC Methylase in 1X GpC Reaction Buffer w/ 600 µM SAM. Incubate reaction mixtures at 37°C.|
|Inactivation||Heat-inactivate the enzyme at 65°C for 20 minutes.|
|Standard Reaction Setup||The reaction volumes can be adjusted accordingly depending on experimental requirements (see protocol).|
DNA sequence after bisulfite treatment. Bisulfite treatment converts non-methylated cytosine to uracil in DNA, which is detected as thymine upon sequencing. Methylated cytosines remain unconverted during treatment. As shown above, treatment of DNA with GpC Methylase methylates cytosines in a GpC context.
The GpC Methylase from Zymo Research catalyzes complete methylation of the GpC sites in DNA. Methylase activity of GpC Methylase was tested for complete methylation of ? DNA using recommended reaction conditions. Completion of GpC methylation was assessed by resistance to digestion with a methylation-sensitive endonuclease (HaeIII) and subsequently analyzed in an agarose gel. "M" is a 1 kb DNA ladder (Zymo Research)