| Recognition site | GGTAC↑C
C↓CATGG |
| Source | An E.coli strain that carries the cloned Kpn I gene from Klebsiella pneumonia |
| 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 | B (10 mM Tris-HCl (pH 7.6 at 25°C); 10 mM MgCl2; 1 mM DTT.) + BSA |
| Enzyme activity (%) | | B | G | O | W | Y | Rose | | 100 | 25 - 50 | 25 - 50 | 25 - 50 | 75 - 100 | 50 |
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| Optimal temperature | 37oC |
| Storage conditions | 10 mM Tris-HCl(pH 7.5); 50 mM NaCl; 0,1 mM EDTA; 1 mM DTT; 200 μg/ml BSA; 50% glycerol; Store at -20°C. |
| Ligation | After 20-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 20 u.a. of enzyme for 16 hours at 37°C.
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| Reagents Supplied with Enzyme |
10 X SE-buffer B, BSA
For E079T and E080T an additional 10X SE-buffer ROSE+ is supplied. |
| Methylation sensitivity | Not blocked by overlapping Dcm methylation (CmCWGG): GGTACCWGG |
| Inactivation 20 minutes under | 80oC |
| Notes | High enzyme concentration may result in star activity.
Long incubation with BSA is not recommended due to star activity.
To obtain 100% activity, BSA should be added to the 1 x reaction mix to a final concentration of 100 μg/ml.
Turbo Kpn 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:
Applications:
-Fast DNA analysis
-Fast preparation of vectors for cloning
-Double digestion
Enzyme Properties:
1 μl of Turbo Kpn I cuts 1 μg of DNA in 1 x SE-Buffer B with BSA 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 (E079).
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 Kpn I
Incubate at 37°C for 15 min.
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| Quality control | Restriction Endonucleases: Quality Control |
| MSDS: | Download MSDS as PDF |
| References: | Smith, D.I., Blattner, F.R., Davies, J. Nucleic Acids Res. 3: 343-353 (1976).
 Abdurashitov M.A., Tomilov V.N., Chernukhin V.A., Gonchar D. A., Degtyarev S. Kh. Comparative analysis of human chromosomal DNA digestion with restriction endonucleases in vitro and in silico. // "Medical genetics" V.6, No 8, pp 29-36, 2007
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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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