Recognition Sequence :
↑(N)7GAACNNNNNNTAC(N)12↑
↓(N)12CTTGNNNNNNATG(N)7↓
Unit Definition :
One unit of the enzyme is the amount required to hydrolyze 1 μg of T7 DNA in 1 hour at 30°C in a total reaction volume of 50 μl.
Reaction Temperature :
37°C
Storage Buffer :
10 mM Tris-HCl (pH 7.5); 50 mM KCl; 0,1 mM EDTA; 7 mM 2-mercaptoethanol; 200 μg/ml BSA, 50% glycerol.
Ligation :
After 2-fold overdigestion with enzyme more than 70% of DNA fragments can be ligated. Of these 80% can be recut. In the presence of 10% PEG ligation is better.
Source :
Pseudomonas stutzeri N2
Working buffer :
Y (33 mM Tris-acetate (pH 7.9 at 25°C); 10 mM magnesium acetate; 66 mM potassium acetate; 1 mM DTT.) + BSA
Non-specific hydrolisis :
No nonspecific activity was detected after incubation of 1 μg of T7 DNA with 2 u.a. of enzyme for 16 hours at 30°C.
Concentration, u.a./ml :
1000-2000
Inactivation :
20min Under 65°C
Reagents Supplied :
10 X SE-buffer Y, BSA
Notes :
High enzyme concentration may result in star activity.
Incubation at 37centigrade results in 20% activity.
To 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.
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