The enzyme cleaves C5-methylated DNA and doesn't cut unmodified DNA.
10 mM Tris-HCl (pH 7.5); 200 mM NaCl; 0,1 mM EDTA; 7 mM 2-mercaptoethanol; 200 µg/ml BSA; 50% glycerol.
Incubate at 55°C.
1XSEBuffer W (pH 8.5 @ 25ºC)
10 mM Tris-HCl,100 mM NaCl, 10 mM MgCl2,1 mM DTT
One unit is defined as the amount of enzyme required to hydrolyze completely 1 μg of linearized plasmid
pHspAI10/DriI+M.Fsp4HI in 1 hour at 55°C in a total reaction volume of 50μl. pHspAI10/DriI+M.Fsp4HI is a plasmid pHspAI10, which is linearized with DriI, and, additionally, modified with Fsp4HI DNA methyltransferase pHspAI10 carries a
gene of HspAI DNA methyltransferase, that modifies the sequence . 5`-GCGC-3`, producing 5`-G(5mC)GC-3`. M.Fsp4HI modifies the sequence 5`-GCNGC-3`, producing 5`-G(5mC)NGC-3`. A substrate
pHspAI10/DriI+M.Fsp4HI includes one site
5`G(5mC)G(5mC)NG(5mC)G(5mC)-3`/3`(5mC)G(5mC)GN(5mC)G(5mC)G-5`,which is MteI canonical site . The enzyme activity depends on a number and positions of methylated nucleotides in the recognition sequence. For example, MteI cuts the
recognition site with six 5-methylcytosines, but the enzyme activity is reduced for more that one order 
Quality Control Assays
No detectable degradation of 1μg of Lambda DNA was observed after incubation with 10 units of enzyme for 16 hours at 55°C in a total reaction volume
of 50 μl.
No detectable degradation of a single- and double-stranded oligonucleotide was observed after incubation with 10 units of enzyme for 3 hours.
Activity in SEBuffers:
SEBuffer B 25- 50%
SEBuffer G 75-100%
SEBuffer O 75-100%
SEBuffer W 100%
SEBuffer Y 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 a complete digestion.
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