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You are here:Home » Molecular Biology » MB-Enzymes, Restriction » PaeI (SphI)

PaeI (SphI)

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Specifications

Catalog #P1800
Sequence5'-G C A T G^C-3'
3'-C^G T A C G-5'
Source Pseudomonas aeruginosa
Concentration 10u/ul
Unit DefinitionOne unit is defined as the amount of PaeI required to digest 1ug of lambda DNA fragments in 1 hr at 37°C in 50ul assay buffer.
Diluent BufferSupplied as a liquid in 10mM Tris-HCl, pH 7.4, 100mM KCl, 1mM EDTA, 1mM DTT, 0.2mg/ml BSA, 50% glycerol.
Storage Buffer10mM potassium phosphate (pH 7.0 at 25°C), 50mM KCl, 1mM DTT, 0.1mM EDTA, 0.5mg/ml BSA, 0.15% Triton X-100 and 50% glycerol.
Supplied withR1625: Supplied as a liquid in 33mM Tris-acetate, 10mM magnesium acetate, 66mM potassium acetate, 0.1mg/ml BSA, pH 7.9 at 37ºC.
R1625-01 Restriction Enzyme Buffer B, 10X: Supplied as a liquid 10mM Tris-HCl, pH 7.5 at 37°C, 10mM magnesium chloride, 0.1mg/ml BSA. Incubate at 37°C
Double DigestsR1625 is provided to simplify buffer selection for double digests. 98% of PaeI, works well in a 1x or 2x.
Overdigestion Assay
No detectable change in the specific fragmentation pattern is observed after 160-fold overdigestion (10u/ug lambda DNA x 16 hours) with PaeI.
Ligation/Recutting Assay
After 50-fold overdigestion (3u/ug DNA x 17 hours) with PaeI, more than 95% of the DNA fragments can be ligated at a 5'-termini concentration of 0.06uM. More than 95% of these can be recut.
Labeled Oligonucleotide (LO) Assay
No detectable degradation of a single-stranded and double-stranded labeled oligonucleotide was observed after incubation with 10 units of PaeI for 4 hours.
Blue/White Cloning Assay
pUC57 was incubated with 10 units of PaeI for 16 hours. After religation and transformation <1% of white colonies were detected.
Methylation Effects
Dam, Dcm, EcoKI: never overlaps-no effect
CpGMay overlap- no effect
EcoBIMay overlap- blocked
Stability during Prolonged Incubation
A minimum of 0.1 units of PaeI is required for complete digestion of 1ug of lambda DNA in 16 hours at 37°C.
Thermal Inactivation
PaeI is inactivated by incubation at 65°C for 20min.
Digestion of Agarose-embedded DNA
A minimum of 5 units of PaeI is required for digestion of 1ug of agarose-embedded lambda DNA in 16 hours.
Number of Recognition Sites in DNA
Lambda 6
pBR322 1
phiX174 0
pUC57, pUC18/19, pTZ19R/U, M13mp18/19 1


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