|Source||An E.coli strain that carries the cloned Sph I gene from Streptomyces phaeochromogenes|
|Assayed on||Lambda DNA|
|Unit definition||One 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.|
|Optimal SE-buffer||G (10 mM Tris-HCl (pH 7.6 at 25°C); 10 mM MgCl2; 50 mM NaCl; 1 mM DTT.) + BSA|
|Enzyme activity (%)|
|25 - 50||100||75 - 100||75 - 100||50 - 75||100|
|Storage conditions||10 mM Tris-HCl (pH 7.5); 100 mM NaCl; 0,1 mM EDTA; 1 mM DTT; 100 μg/ml BSA; 50% glycerol. Store at -20°C.|
|Ligation||After 10-fold overdigestion with enzyme more than 90% of the DNA fragments can be ligated and recut.|
|Non-specific hydrolisis||No nonspecific activity was detected after incubation of 1 μg of Lambda DNA with 10 u.a. of enzyme for 16 hours at 37°C.
|Reagents Supplied with Enzyme||
10 X SE-buffer G, BSA (except E129T and E130T).|
For E129T and E130T an additional 10X SE-buffer ROSE is supplied.
|Methylation sensitivity||not tested|
|Inactivation 20 minutes under||65oC|
|Notes||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.
Turbo Sph I can be used for short time (15 min) DNA digestion as well as for standard reaction. The reaction can be performed using optimal or universal (ROSE) Buffer.
Turbo DNA Digestion:
-Fast DNA analysis
-Fast preparation of vectors for cloning
1 μl of Turbo Sph I cuts 1 μg of DNA in 1 x SE-Buffer G or universal 1 x SE-Buffer ROSE in 15 min (see the protocol below). A short time of DNA digestion requires high quality purification of DNA sample. The enzyme may be used in standard conditions of DNA cleavage (E129).
Turbo reaction protocol:
20 μl of the reaction volume:
10 x SE Buffer ROSE - 2 μl
DNA - 0,2-1 μg
Nuclease-free water - to 20 μl
+ 1μl of Turbo Sph I
Incubate at 37°C for 15 min.
|Quality control||Restriction Endonucleases: Quality Control|
|MSDS:||Download MSDS as PDF|
|References:||Fuchs, L.Y., Covarrubias, L., Escalante, L., Sanchez, S., Bolivar, F. Gene 10:39-46 (1980).|
|Theoretical diagrams of DNA digestion by this enzyme for the most known DNA substrates:|
To view the fragments length values please point mouse cursor over diagram
M - ladder, 1 - Adeno-2 DNA, 2 - Lambda DNA, 3 - T7 DNA, 4 - pUC19, 5 - pBR322