| Recognition site | AGC↑GCT
TCG↓CGA |
| Source | E.coli strain that carries the cloned Afe I gene from Alcaligenes faecalis T2774 |
| Assayed on | Lambda DNA (BamHI-digest) |
| Unit | One unit of the enzyme is the amount required to hydrolyze 1 μg of Lambda DNA (BamHI-digest) in 1 hour at 37°C in a total reaction volume of 50 μl. |
| Optimal SE-buffer | Y (33 mM Tris-acetate (pH 7.9 at 25°C); 10 mM magnesium acetate; 66 mM potassium acetate; 1 mM DTT.) |
| Enzyme activity (%) | | B | G | O | W | Y | R | | 10 - 25 | 25 - 50 | 75 - 100 | 75 - 100 | 100 | 100 |
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| Optimal temperature | 37oC |
| Storage conditions | 10 mM Tris-HCl (pH 7.6); 50 mM NaCl; 0.1 mM EDTA; 1 mM DTT; 200 μg/ml BSA; 50% glycerol; Store at -20°C. |
| Ligations | After 10-fold overdigestion with enzyme more than 80% of DNA pBR322 fragments can be ligated and recut. |
| Non-specific hydrolisis | No nonspecific activity was detected after incubation of 1 μg of 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 Y |
| Methylation sensitivity | not tested |
| Inactivation 20 minutes under | 65oC |
| Notes | The minimum number of units that resulted in complete digestion of 1 μg of substrate DNA in 16 hours is 0,25.
AfeI cleaves supercoiled and linear plasmid DNA (pBR322) at a roughly equal rate. AfeI cleaves Lambda DNA/BamHI digest at a rate 3-4 times higher than plasmid DNA. |
| MSDS: | Download MSDS as PDF |
| References: | Abdurashitov, M.A., Kileva, E.V., Shevchenko, A.V., Degtyarev, S.Kh.
Unpublished observations. (1994)
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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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