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You are here:Home » Molecular Biology » MB-Enzymes, Restriction » Eam1104I (Ksp632I)

Eam1104I (Ksp632I)

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Specifications

5'-C T C T T C(N)1^-3'
3'-G A G A A G(N)4^-5'
Catalog #E0200
Concentration 10u/ul.
SourceEnterobacter amnigenus RFL1104
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 Temperature37°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 as a liquid in10mM Tris-HCl (pH 7.4 at 25°C), 100mM KCl, 1mM DTT, 1mM EDTA, 0.2mg/ml BSA, 50% glycerol.
Supplied WithR1625: Restriction Enzyme Buffer A, 10X. Provided to simply buffer selection.
Storage and StabilityMay be stored at 4°C for short-term only. For long-term storage, aliquot and store at -20°C. Aliquots are stable for at least 6 months at -20°C. For maximum recovery of product, centrifuge the original vial after thawing and prior to removing the cap. Further dilutions can be made in assay buffer.
Quality Control
Overdigestion AssayNo detectable change in the specific fragmentation pattern is observed after 320-fold overdigestion (20u/ug lambda DNA x 16 hours) with Eam1104I
Ligation/Recutting Assay
After 50-fold overdigestion (3u/ug DNA x 17 hours) with Eam1104I, more than 95% of the DNA fragments can be ligated at a 5'-termini concentration of 0.2uM. More than 95% 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 rEam1104I for 4 hours.
Enzyme Properties
Stability during Prolonged Incubation
A minimum of 0.5u of Eam1104I is required for complete digestion of 1ug of lambda DNA in 16 hours at 37°C.
Thermal Inactivation
Eam1104I is inactivated by incubation at 65°C for 20min.
Digestion of Agarose-Embedded DNA
A minimum of 5u of Eam1104I is required for complete digestion of 1ug of agarose-embedded lambda DNA in 16 hours.
Number of Recognition Sites in DNA
Lambda34
PhiX174, M13mp18/19, pBR3222
pUC18/19, pUC57, pTZ19R/U: 3
Protocol for Digestion
Add
Nuclease free water16ul
R16252ul
DNA (0.5-1ug/ml)1ul
E02000.5-2ul
Mix gently and spin down for a few seconds. Incubate at 37ºC for 1-16 hours.
Protocol for Digestion Directly after Amplification
Add
PCR Reaction Mixture10ul (~0.1-0.5ug of DNA)
Nuclease free water18ul
R16252ul
E02001-2ul
Mix gently and spin down for a few seconds.
Incubate at 37ºC for 1-16 hours.


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