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You are here:Home » Molecular Biology » MB-Enzymes, Restriction » BfuI (BciVI)

BfuI (BciVI)

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Specifications

Catalog #B1086-05
Sequence5'-G T A T C C (N)6^-3'
3'-C A T A G G (N)5^-5'
Source Bacillus firmus Auk
Concentration 5u/ul
Unit DefinitionOne unit is defined as the amount of BfuI required to digest 1ug of lambda DNA dcm in 1 hour at 37°C in 50ul of reaction buffer.
Supplied WithR1625: Restriction Enzyme Buffer A, 10X: Supplied as a liquid in 33mM Tris-acetate, 10mM magnesium acetate, 66mM potassium acetate, 0.1mg/ml BSA (pH 7.9 at 37°C).
R1625-20 Restriction Enzyme Buffer for Bful, 10X: Supplied as a liquid in 10mM Tris-HCl (pH 7.4 at 25°C), 300mMNaCl, 1mM DTT, 1mM EDTA, 0.2mg/ml BSA, 50% glycerol.
Storage and Dilution Buffer10mM Tris-HCl (pH 7.4 at 25ºC), 100mM KCl, 1mM EDTA, 1mM DTT, 0.2mg/ml BSA, 50% glycerol.
Thermal InactivationBful is inactivated by incubation at 80°C for 20min.
Overdigestion Assay
No detectable change in the specific fragmentation pattern is observed after 5-fold overdigestion (5u/ug lambda DNA x 1 hour) with BfuI.
Ligation/Recutting Assay
After 5-fold overdigestion (5u/ug DNA x 1 hour) with BfuI, more than 80% of the DNA fragments can be ligated in a reaction mixture containing 2040u of T4 DNA ligase/1ug of fragments and 10% PEG at a 5'-termini concentration of 0.25uM. More than 90% 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 10units of BfuI for 4 hours.
Star Activity
A large excess of BfuI (7.5u/ug DNA x 1 hour) may result in star activity.
Stability during Prolonged Incubation
A minimum of 1.0unit of BfuI is required for complete digestion of 1ug of lambda DNA in 16 hours at 37°C.
Digestion of Agarose-embedded DNA
A minimum of 5units of BfuI is required for complete digestion of 1ug of agarose-embedded lambda DNA in 16 hours.
Recommended Protocol for Digestion
Add
Nuclease free water16ul
R1625-202ul
DNA (0.5-1ug/ul)1ul
Bful0.5-2ul
Mix gently, spin down for a few seconds.
Incubate at 37ºC for 1-16 hours.
Protocol for Digestion of PCR products directly after amplification
Add
PCR Reaction Mixture10ul (~0.1-0.5ug of DNA)
Nuclease free water18ul
R1625-202ul
Bful1-2ul
Mix gently, spin down for a few seconds.
Incubate at 37ºC for 1-16 hours.


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