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You are here:Home » Molecular Biology » MB-Enzymes, Restriction » BoxI (PshAI)

BoxI (PshAI)

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Specifications

5'-G A C N N^N N G T C-3'
3'-C T G N N^N N C A G-5'
Catalog #B2620
Concentration 10u/ul
Source Brevibacterium oxydans Iti 12–052
Buffer 33mM Tris-acetate (pH 7.9), 10mM magnesium acetate, 66mM potassium acetate and 0.1mg/ml BSA. Incubate at 37°C. (Incubation at 30°C results in a 2.5-fold increase in activity.)
Diluent Buffer 10mM Tris-HCl (pH 7.4 at 25°C), 100mM KCl, 1mM EDTA, 1mM DTT, 0.2mg/ml BSA and 50% glycerol. For longer periods–the Storage Buffer should be used.
Storage Buffer 10mM potassium phosphate (pH 7.4 at 25°C), 100mM NaCl, 1mM EDTA, 7mM 2-mercaptoethanol, 0.2mg/ml BSA and 50% glycerol.
Overdigestion AssayNo detectable change in the specific fragmentation pattern is observed after 160-fold overdigestion (10u/ug lambda DNA x 16 hours) with BoxI.
Ligation/Recutting AssayAfter 50-fold overdigestion (3u/ug DNA x 17 hours) with BoxI, more than 90% of the DNA fragments can be ligated at a 5'-termini concentration of 0.08uM. More than 95% of these can be recut.
Labeled Oligonucleotide (LO) AssayNo detectable degradation of a single-stranded and double-stranded labeled oligonucleotide was observed after incubation with 10units of restriction endonuclease for 4 hours.
Blue/White Cloning Assay The mix of pUC57/PstI, pUC57/Eco32I and pUC57/HindIII digests was incubated with 10units of enzyme for 5 hours. After religation and transformation 0.2% of white colonies were detected.
Methylation EffectsThe hemi-methylated sequence GAm5C(N)4GTC is cleaved by BoxI at a significantly slower rate. Cleavage is blocked when both strands are methylated as shown: 5'-GAm5C GNNNNm5C GTC-3' 3'-CT Gm5CNNNN Gm5CAG-5' Overlapping CG methylation may influence DNA cleavage.
Stability during Prolonged IncubationA minimum of 0.5units of enzyme is required for complete digestion of 1ug of DNA in 16 hours at 37°C.
Thermal InactivationEnzyme is inactivated by incubation at 80°C for 20min.
Digestion of Agarose-embedded DNAA minimum of 10units of enzyme is required for digestion of 1ug of agarose-embedded lambda DNA in 16 hours.
Number of Recognition Sites in DNA
Lambda7
PhiX1741
M13mp18/190
pBR3221
pUC18/190
pUC570
pTZ19R/U0
pBluescriptIIKS(-/+)0
pBluescriptIISK(-/+)0
pACYC1770
pACYC1842
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.


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