BstAF I

Enzyme nameBstAF I
PrototypeAflII
Known analogsAflII, BfrI, BspTI, Bst98I, MspCI, Vha464I
SKUSE-E135
Turbo versionNot available
High-concentration versionNot available
Restriction typeType IIPiType IIP enzymes are classical Type II restriction enzymes with palindromic recognition sequences and symmetric cleavage sites.
Recognition site
5'… C▼TTAAG …3'
3'… GAATT▲C …5'
SourceBacillus stearothermophilus AF
Optimal bufferSE-buffer W (10 mM Tris-HCl (pH 8.5 at 25°C); 10 mM MgCl2; 100 mM NaCl; 1 mM DTT.)
Optimal temperature55 °C
Inactivation temperature80 °C
Buffer activity
BGOWYROSE
10257510025100
Unit definitionOne unit of the enzyme is the amount required to hydrolyze 1 μg of Lambda DNA in 1 hour at 55°C in a total reaction volume of 50 μl.
Assayed onLambda DNA
Storage conditions10 mM Tris-HCl (pH 7.5); 200 mM KCl; 0.1 mM EDTA; 7 mM 2-mercaptoethanol; 200 μg/ml BSA; 50% glycerol. Store at -20°C.
LigationAfter 5-fold overdigestion with enzyme ~40% of the DNA fragments can be ligated and 95% of these can be recut. In the presence of 10% PEG
Nonspecific hydrolysisNo nonspecific activity was detected after incubation of 1 μg of Lambda DNA with 40 u.a. of enzyme for 16 hours at 55°C.
Methylation sensitivitynot tested
Reagents supplied with enzyme10 X SE-buffer W
NotesDo not use BSA for long incubation. To obtain 100% activity, BSA should be added to the 1 x reaction mix to a final concentration of 100 μg/ml.
References

No publications found.

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