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You are here:Home » Molecular Biology » MB-Enzymes, Restriction » AdeI (DraIII)

AdeI (DraIII)

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Specifications

5'-C A C N N N^G T G-3'
3'-G T G^N N N C A C-5'
Catalog #A0859-75
SourceAlcaligenes denitrificans SS 3028
Concentration 10u/ul
Unit Definition:
One unit is defined as the amount of AdeI required to digest 1ug of lambda DNA in 1 hour at 37°C in 50ul of assay buffer.
Incubation Temperature 37°C
Diluent Buffer10mM Tris-HCl (pH 7.4 at 25°C), 100mM KCl, 1mM EDTA, 1mM DTT, 0.2mg/ml BSA and 50% glycerol.
Storage BufferSupplied as a liquid in 10mM Tris-HCl (pH 7.5 at 25°C), 300mM KCl, 1mM DTT, 0.1mM EDTA, 0.2mg/ml BSA and 50% glycerol.
Supplied withR1625-02: Restriction Enzyme Buffer C, 10X:
Supplied as a liquid in 100mM Tris-HCl (pH 7.5), 100mM magnesium chloride, 500mM sodium chloride, 1mg/ml BSA..
R1625 Restriction Enzyme Buffer A, 10X:
Supplied as a liquid in 330mM Tris-acetate (pH 7.9) 100mM magnesium acetate, 660mM potassium acetate, 1mg/ml BSA.
Enzyme PropertiesStar Activity:
An excess of AdeI (20u/µg DNA x 1 hour) may result in star activity.
Methylation EffectsDam, Dcm: Never overlaps - No effect.
CpG: May overlap - cleavage impaired.
EcoKl, EcoBl: May overlap - Effect not determined.
Stability during Prolonged IncubationA minimum of 0.5units of AdeI is required for complete digestion of 1ug of DNA in 16 hours at 37°C.
Digestion of Agarose-embedded DNAA minimum of 10units of AdeI is required for complete digestion of 1ug of agarose- embedded lambda DNA in 16 hours.
Thermal InactivationAdeI is not inactivated by incubation at 80°C for 20min (see Protocol for Inactivation Procedure.
Number of Recognition Sites in DNA:
Lambda: 10
PhiX174, pTZ19R/U, M13mp18/19, pBluescriptIIKS(-/+), pBluescriptIISK(-/+), pACYC177: 1
pBR322, pUC18/19, pUC57, pACYC184: 0
Quality Control
Overdigestion Assay
No detectable change in the specific fragmentation pattern is observed after 15-fold overdigestion (15u/ug lambda DNA x 1hour) with AdeI (see Star Activity).
Ligation and Recleavage (L/R) Assay
The L/R 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 (LO) Assay
No detectable degradation of a single-stranded and double-stranded labeled oligonucleotide was observed after incubation with 10 units of AdeI for 4 hours.
Blue/White (B/W) Cloning Assay
The B/W assay was replaced with LO tests after validating experiments showed LO test ability to detect nuclease and phosphatase activities with sensitivity that equals to that of B/W test.
Storage and Stability
May be stored at 4°C for short-term only. Aliquot to avoid repeated freezing and thawing. Store at -20°C. Aliquots are stable for 6 months at -20°C. For maximum recovery of product, centrifuge the original vial after thawing and prior to removing the cap.


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