Supplied in:

20 mM Tris-HCl (pH 7.4); 200 mM NaCl; 0,1 mM EDTA; 7 mM  2-mercaptoethanol; 50% glycerol.

Reaction Conditions:

1X SEBuffer Y. 

Incubate at  37°C.

1X SEBuffer Y  (pH 7.9 @ 25ºC)

33 mM Tris-Ac, 66 mM KAc, 10 mM MgAc, 1 mM DTT

Unit Definition:

One unit is defined as the amount of enzyme required to hydrolyze at least one of three canonical sites: 5`-G(5mC)NG(5mC)-3`/3`-CGN(5mC)G-5` in 1 μg of linearized plasmid pFsp4HI3 in 1 hour at 37°C in a total reaction volume of 50 μl. DNA pFsp4HI3/DriI is a linearized plasmid pFsp4HI3, which carries a gene of DNA-methyltransferase M.Fsp4HI and includes  three canonical sites: 5`-G(5mC)NG(5mC)-3`/3`-CGN(5mC)G-5` [2].

Quality Control Assays

16-Hour Incubation:

No detectable degradation of 1μg of Lambda DNA was observed after incubation with 2 units of enzyme for 16 hours at 37°C in a total reaction volume of 50 μl.

Oligonucleotide Assay:

No detectable degradation of a single- and double-stranded oligonucleotide was observed after incubation with 2 units of enzyme for 3 hours.

Enzyme Properties

Activity in SEBuffers:

SEBuffer B      50-75%

SEBuffer G     75-100%

SEBuffer O      10-25%

SEBuffer W     25-50%

SEBuffer Y         100%

SEBuffer ROSE  100%

When using a buffer other than the optimal (suppied) SEBuffer, it may be necessary to add more enzyme to achieve a complete digestion.

Reagents supplied with enzyme:

10X SEBuffer Y

Heat Inactivation:

Yes   (65°C for 20 minutes)