|Molecular Biology||Storage: -20°CShipping: Blue Ice|
5'-T^G A T C A-3'
3'-A C T A G^T-5'
Source: Bacillus caldolyticus
Buffer: 10mM Tris-HCl (pH 7.5), 10mM MgCl2, 50mM NaCl and 0.1mg/ml BSA. Incubate at 55°C. Incubation at 37°C results in 50% activity.
Double Digests Y+/Tango™ Buffer is provided to simplify buffer selection for double digests. 98% of Fermentas Restriction Endonucleases (REs) work well in a 1X or 2X concentration of the Y+/Tango™ Buffer. Please refer to the Enzyme Activity Chart to choose the best buffer for the two REs in your digest.
Diluent Buffer: 10mM Tris-HCl (pH 7.4 at 25°C), 100mM KCl, 1mM EDTA, 1mM DTT, 0.2mg/ml BSA and 50% glycerol.
Storage Buffer: 10mM Tris-HCl (pH 7.4 at 25°C), 100mM KCl, 1mM DTT, 1mM EDTA, 0.2mg/ml BSA and 50% glycerol.
Overdigestion Assay: No detectable change in the specific fragmentation pattern is observed after 80-fold overdigestion (5u/ug lambda DNA x 16 hours) with BclI (see Star Activity).
Ligation/Recutting Assay: After 50-fold overdigestion (3u/ug DNA x 17 hours) with BclI, more than 90% of the DNA fragments can be ligated at a 5'-termini concentration of 0.1uM. 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 restriction endonuclease for 4 hours.
Blue/White Cloning Assay: The mix of pUC57/HindIII, pUC57/PstI and pUC57/Eco32I digests was incubated with 10units of enzyme for 16 hours. After religation and transformation 0.3% of white colonies were detected.
Star Activity: A large excess of enzyme (7.5u/ug DNA x 16 hours) may result in star activity. Star activity appears at 10-fold overdigestion (10 units x 1 hour).
Methylation Effects: BclI cuts neither TGm6ATCA nor TGAThm5CA, but cuts TGATm5CA.
Blocked by Dam methylation.
Stability during Prolonged Incubation: A minimum of 0.1unit of enzyme is required for complete digestion of 1ug of lambda DNA in 16 hours at 55°C.
Thermal Inactivation: Only small amounts of enzyme (up to 10units) can be inactivated at 80°C for 20min.
Compatible Ends: BamHI, BglII, Bsp143I, MboI, PsuI
Number of Recognition Sites in DNA:
Unit Definition: One unit is defined as the amount of enzyme required to digest 1ug of lambda DNA dam-in 1 hour at 55°C in 50ul of assay buffer.