| Recognition site | ATTT↑AAAT
TAAA↓TTTA |
| Source | Streptococcus milleri S |
| Assayed on | T7 DNA (SspI-digest) |
| Unit definition | One unit of the enzyme is the amount required to hydrolyze 1 μg of T7 DNA (SspI-digest) in 1 hour at 37°C in a total reaction volume of 50 μl. |
| Optimal SE-buffer | O (50 mM Tris-HCl (pH 7.6 at 25°C); 10 mM MgCl2; 100 mM NaCl; 1 mM DTT.) + BSA |
| Enzyme activity (%) | | B | G | O | W | Y | Rose | | 25 - 50 | 25 - 50 | 100 | 75 - 100 | 25 - 50 | 10 |
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| Optimal temperature | 37oC |
| Storage conditions | 10 mM Tris-HCl (pH 7.5); 250 mM NaCl; 0.1 mM EDTA; 7 mM 2-mercaptoethanol; 100 μg/ml BSA; 50% glycerol. Store at -20°C. |
| Ligation | After 20-fold overdigestion with enzyme >95% of the DNA fragments can be ligated and recut. Ligation >95% in presence of 10% PEG. |
| Non-specific hydrolisis | No nonspecific activity was detected after incubation of 1 μg of T7 DNA with 40 u.a. of enzyme for 16 hours at 37°C.
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| Reagents Supplied with Enzyme |
10 X SE-buffer O, BSA
For E225T and E226T 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 Smi I can be used for short time (10 min) DNA digestion as well as for standard reaction. The reaction can be performed using optimal or universal (ROSE) Buffer
Turbo DNA Digestion:
Applications:
-Fast DNA analysis
-Fast preparation of vectors for cloning
-Double digestion
Enzyme Properties:
1 μl of Turbo Smi I cuts 1 μg of DNA in 1 x SE-Buffer O or universal 1 x SE-Buffer ROSE in 10 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 (E225).
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 Smi I
Incubate at 37°C for 10 min.
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| Quality control | Restriction Endonucleases: Quality Control |
| MSDS: | Download MSDS as PDF |
| References: | Dedkov, V.S., Bondar, T.S., Shevchenco, A.V., Degtyarev, S.Kh. Mol. Gen. Mikrobiol. Virusol. 1: 23-27 (2000).
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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
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