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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
Supplied WithR1625: Restriction Enzyme Buffer A, 10X
R1625-03: Restriction Enzyme Buffer D, 10X
Unit Definition One 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
DilutionFor short-term storage (3-4 weeks):
Dilute with Diluent Buffer (R1625-05, 10mM Tris-HCl, pH 7.4 at 25°C, 100mM potassium chloride, 1mM EDTA, 1mM DTT, 0.2mg/ml BSA, 50% glycerol.
For long-term storageDilute with Storage Buffer (10mM potassium phosphate, pH 7.4, 25°C, 300mM NaCl, 1mM EDTA, 7mM 2-mercaptoethanol, 0.2mg/ml BSA, 0.15% Triton X-100, 50% glycerol).
Quality ControlOverdigestion Assay: No 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 After 50-fold overdigestion (3u/ug DNA x 17 hours) with EcoRI, more than 90% of the DNA fragments can be ligated at a 5'-termini concentration of 0.02uM. More than 90% of these can be recut.
Labeled Oligonucleotide Assay No detectable degradation of a single-stranded and double-stranded labeled oligonucleotide was observed after incubation with 10u of restriction endonuclease for 4 hours.
Blue/White Cloning Assay pUC57 was digested at a unique site with 10u of enzyme for 16 hours. After religation and transformation 0.2% of white colonies were detected.
Methylation Effects EcoRI cuts neither Gm6AATTC nor GAm6ATTC, but cuts GAATTm5C, GAATThm5C and GAAhm5Uhm5UC.
Storage and StabilityStore at -20C. Stable for 12-12 months.
SourceE.coli that carries the cloned ecoRIR gene from Escherichia coli RY13
Concentration10u/ul.
(See E0375-05 for 50u/ul).
Enzyme Properties
Star ActivityStar Activity appears at 10-fold overdigestion (10u x 1 hour). A large excess of the enzyme (10u/ug DNA x 16 hours), low salt concentration, high pH, or the replacement of Mg2+ by Mn2+ results in star activity.
Stability during Prolonged IncubationA minimum of 0.2u of enzyme are required for complete digestion of 1ug of lambda DNA in 16 hours at 37°C.
Thermal InactivationEnzyme is inactivated by incubation at 65°C for 20min.
Digestion of Agarose-Embedded DNAA minimum of 5u of enzyme 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
Important NoteThis product as supplied is intended for research use only, not for use in human, therapeutic or diagnostic applications without the expressed written authorization of United States Biological.


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