10 mMKH2PO4 (pH
7.45); 200 mM NaCl; 0,1 mM EDTA;200µg/ml
2-mercaptoethanol; 50% glycerol.
Incubate at 37°C.
Y(pH 7.9 @ 25ºC)
66 mMKAc 1 mM DTT
The enzyme activity depends on number and position of methylated cytosines.
Optimal recognition site:
Unit Definition:One unit is defined as the amount of enzyme required to hydrolyzecompletely a unique site 5`-G(5mC)NG(5mC)-3`/3`-(5mC)GN(5mC)G-5` in 1μg oflinearized plasmid pFsp4HI3/DriI in 1 hour at 37°C in a total reaction volume of 50μl.
DNA pFsp4HI3/DriIis a linearized plasmid pFsp4HI3, whichcarriesa gene
of DNA-methyltransferase M.Fsp4HI and includes a unique GluI site:
Quality Control Assays
No detectable degradation of 1μg of Lambda DNA was observed after incubation with 1 units of enzyme
for 16 hours at 37°C in a total reaction volume of 50μl.
No detectable degradation of a single- and double-stranded oligonucleotide was
observed after incubation with 1 units of enzymefor 3 hours.
SEBuffer B 75-100%
SEBuffer G 75-100%
SEBuffer O 25-50%
SEBuffer W 50-75%
SEBuffer ROSE 100%
When using a buffer other than the optimal (suppied) SEBuffer, it may be
necessary to add more enzyme to achieve complete digestion.