Description: MspI Methyltransferase recognizes the same sequence as the HpaII Methyltransferase, but modifies the external cytosine residue (C5) of the sequence CCGG.
Source: An E. coli strain that carries the cloned MspI modification gene from Moraxella species (ATCC 49670).
Reaction Conditions: 1X MspI Methylase Reaction Buffer Supplemented with 80 μM S-adenosylmethionine (SAM) Incubate at 37°C.
1X MspI Methylase Reaction Buffer: 50 mM Tris-HCl
100 mM NaCl
5 mM 2-Mercaptoethanol
10 mM EDTA
pH 7.5 @ 25°C
Unit Definition: One unit is defined as the amount of enzyme required to protect 1 µg λ DNA in 1 hour at 37°C in a total reaction volume of 10 µl against cleavage by MspI restriction endonuclease.
Protection Assay Conditions: MspI Methyltransferase is incubated with 1 µg of λ DNA in 10 µl 1X MspI Methyltransferase Reaction Buffer, supplemented with 80 µM S-adenosylmethionine, for one hour at 37°C followed by 15 minutes at 65°C. The extent of protection by MspI Methyltransferase is determined by the addition of 40 µl NEBuffer 2 supplemented with 10 mM MgCl2 and 5 units of MspI restriction endonuclease. Incubation for MspI at 37°C is followed by analysis on agarose gels.
Concentration: 5,000 units/ml
Storage Conditions: 50 mM Tris-HCl
50 mM NaCl
5 mM 2-Mercaptoethanol
10 mM EDTA
200 µg/ml BSA
50% Glycerol
pH 7.5 @ 25°C
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