Pps I

Enzyme namePps I
PrototypePleI
Known analogsPleI, MlyI, SchI
SKUSE-E269
Turbo versionNot available
High-concentration versionNot available
Restriction typeType IISiType IIS enzymes recognize asymmetric DNA sequences and cleave outside the recognition site at a defined distance. They are widely used in Golden Gate assembly.
Recognition site
5'… GAGTC(N)4▼ …3'
3'… CTCAG(N)5▲ …5'
SourcePseudomonas pseudoalcaligenes
Optimal bufferSE-buffer Y (33 mM Tris-acetate (pH 7.9 at 25°C); 10 mM magnesium acetate; 66 mM potassium acetate; 1 mM DTT.)
Optimal temperature37 °C
Inactivation temperature65 °C
Buffer activity
BGOWYROSE
501002510050
Unit definitionOne unit of the enzyme is the amount required to hydrolyze 1 μg of Lambda DNA in 1 hour at 37°C in a total reaction volume of 50 μl.
Assayed onLambda DNA
Storage conditions10 mM Tris-HCl (pH 7.5); 50 mM KCl; 0.1 mM EDTA; 7 mM 2-mercaptoethanol; 200 μg/ml BSA and 50% glycerol; Store at -20°C.
LigationAfter 2-fold overdigestion with enzyme 20% of the DNA fragments can be ligated and recut.
Nonspecific hydrolysisNo nonspecific activity was detected after incubation of 1 μg of Lambda DNA with 1 u.a. of enzyme for 16 hours at 37°C.
Methylation sensitivitynot tested
Reagents supplied with enzyme10 X SE-buffer Y, BSA
NotesTo obtain 100% activity, BSA should be added to the 1 x reaction mix to a final concentration of 100 μg/ml. Do not use BSA for long incubation.
References
  1. Unpublished data Dedkov, V.S., Bondar, T.S., Shevchenko, A.V., Abdurashitov, M.A., Degtyarev, S.Kh Unpublished observations (1998).
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