Properties & Usage
Unit Definition
One unit is defined as the amount of enzyme required to protect 1 µg of λ DNA in a total reaction volume of 20 µl in 1 hour at 37°C against cleavage by HaeIII restriction endonuclease. Reaction Conditions
1X GC Reaction Buffer
Supplement with 640 µM S-adenosylmethionine (SAM)
Incubate at 37°C
1X GC Reaction Buffer
50 mM NaCl
50 mM Tris-HCl
10 mM DTT
(pH 8.5 @ 25°C)
Control Protection Assay
One unit of GpC Methyltransferase is incubated with 1 µg of λ DNA in 20 µl 1X GC Reaction Buffer, supplemented with 640 µM S-adenosylmethionine, for one hour at 37°C followed by 15 minutes inactivation at 65°C . The extent of protection by GpC Methyltransferase is determined by the addition of 10 units of HaeIII restriction endonuclease in 30 µl 1X NEBuffer 2 followed by incubation at 37°C for one hour. Results are analyzed on agarose gel. Storage Buffer
15 mM Tris-HCl
0.2 M NaCl
1 mM DTT
0.1 mM EDTA
200 µg/ml BSA
50% Glycerol
pH 7.4 @ 25°C
Heat Inactivation
65°C for 15 minutes