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You are here:Home » Molecular Biology » MB-Enzymes, Restriction » EcoRI 10u/ul

EcoRI 10u/ul

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Specifications

5'-G^A A T T C-3'
3'-C T T A A^G-5'
Catalog #E0375
SourceE.coli that carries the cloned ecoRIR gene from Escherichia coli RY13
Supplied WithR1625: Restriction Enzyme Buffer A, 10X: Supplied as a liquid in 33mM Tris-acetate, 10mM magnesium acetate, 66mM potassium acetate, 0.1mg/ml BSA (pH 7.9 at 37°C).
R1625-03: Restriction Enzyme Buffer D, 10X: Supplied in 50mM Tris-HCl, pH 7.5, 10mM MgCl2, 100mM NaCl, 0.02% Triton X-100, 0.1mg/ml BSA.
Concentration 10u/ul.
(See E0375-05 for 50u/ul).
Unit DefinitionOne unit is defined as the amount of enzyme required to digest 1ug of lambda DNA in 1 hour at 37°C in 50ul of assay buffer.
Incubation Temperature 37°C
Dilution Buffer10mM Tris-HCl, pH 7.4 at 25°C, 100mM potassium chloride, 1mM EDTA, 1mM DTT, 0.2mg/ml BSA, 50% glycerol.
Storage BufferSupplied in 10mM potassium phosphate, pH 7.4, 25°C, 300mM NaCl, 1mM EDTA, 1mM DTT, 0.2mg/ml BSA, 0.15% Triton X-100, 50% glycerol.
Enzyme Properties
Stability during Prolonged IncubationA minimum of 0.2u of EcoRI are required for complete digestion of 1ug of lambda DNA in 16 hours at 37°C.
Thermal InactivationEcoRI is inactivated by incubation at 65°C for 20min.
Digestion of Agarose-Embedded DNAA minimum of 5u of EcoRI is required for complete digestion of 1ug of agarose-embedded lambda DNA in 16 hours.
Compatible EndsXapI, MunI, TasI
Number of Recognition Sites in DNA
Lambda5
PhiX1740
pBR322; 1
pUC571
pUC18/191
pTZ19R/U1
M13mp18/191
Quality Control
Overdigestion AssayNo detectable change in the specific fragmentation pattern is observed after 80-fold overdigestion (5u/ug lambda DNA x 16 hours) with EcoRI.
Ligation/Recutting Assay
The ligation and recleavage assay was replaced with LO test after validating experiments showed LO test ability to trace nuclease and phosphatase activities with sensitivity that is higher than L/R by a factor of 100.
Labeled Oligonucleotide AssayNo detectable degradation of a single-stranded and double-stranded labeled oligonucleotide was observed after incubation with 10u of EcoRI for 4 hours.
Blue/White Cloning AssayThe B/W assay was replaced with LO test after validating experiments showed LO test ability to detect nuclease and phosphatase activities with sensitivity that equals to that of B/W test.
Methylation Effects
Dam, Dcm, EcoBl, EcoKlNever overlaps - no effect,
CpGMay overlap - cleavage impaired.


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