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EcoRI
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EcoRI is a restriction enzyme which recognizes the following sequence:
5'...G▼AATTC...3'
3'...CTTAA▲G…5'
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Concentration
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10u/μl
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Reaction condition |
Recommended Buffer: Buffer O+
50mM Tris-HCl (pH 7.5), 10mM MgCl2, 100mM NaCl and 0.1mg/ml
BSA |
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Incubation temperature: 37°C |
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Thermal inactivation: 65°C |
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Source
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Escherichia coli RY 13.
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Frequency of Cutting
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| λ | Ad-2 | ΦX174 | pUC18 | M13mp18 | pBR322 |
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| 5 | 5 | 0 | 1 | 1 | 1 |
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Storage
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-20°C
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Storage
buffer
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10mM potassium phosphate
(pH 7.4 at 25°C), 300mM NaCl, 1mM EDTA, 7mM 2-mercaptoethanol,
0.2mg/ml BSA, 0.15% Triton X-100 and 50% glycerol |
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Ligation
and Recleavage
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After 50-fold overdigestion
with EcoRI, approximately more than 90% of the DNA fragments
can be ligated and recut
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Methylation
Effect
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EcoRI cuts neither Gm6AATTC
nor GAm6ATTC, but cuts GAATTm5C, GAATThm5C andGAAhm5Uhm5UC
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Comment |
Star activity may be occurred with glycerol concentrations >12%, enzyme:DNA ratios >25u/μg, in the absence of NaCl, in the presence of Mn2+, or under high pH conditions. |
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