| 5'-^ 8(N) G A G (N)5 C T C (N)13^-3' |
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| 3'-^13(N) C T C (N)5 G A G (N) 8^-5' |
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| Catalog # | B2645 |
| BplI requires only Mg2+ for its activity, but is stimulated by S-adenosylmethionine. 0.05mM S-adenosylmethionine gives more than a 100-fold increase of BplI activity. The cleavage of DNA by BplI is never complete. Source | Bacillus pumilus |
| Concentration | 5 units/ul |
| Unit Definition | One unit is defined as the amount of Bpl I required to digest 1ug of lambda DNA-Xhol in 1 hour at 37°C in 50ul of assay buffer (a maximal cleavage level is achieved at which no change in the fragmentation pattern is observed with further increase of enzyme). |
| Storage Buffer | Supplied as a liquid in 10mM Tris-HCl, pH 7.5, 100mM KCl, 1mM DTT, 1mM EDTA, 0.2mg/ml BSA, 50% glycerol. |
| Buffers Supplied | R1625 Restriction Enzyme Buffer A, 10X: |
| 33mM Tris-acetate, pH 7, 10mM magnesium acetate, 66mM potassium acetate, 0.1mg/ml BSA |
| B2645-01 SAM |
| 0.05mM S-adenosylmethionine |
| Storage and Stability | Aliquot to avoid repeated freezing and thawing. Store at -20°C. For maximum recovery of product, centrifuge the original vial after thawing and prior to removing the cap. |
| Stability during Prolonged Incubation | |
| A minimum of 0.3 units of Bpl I is required for complete digestion of 1ug of DNA in 16 hours at 37°C. |
|
| Thermal Inactivation | |
| Bpl I is inactivated by incubation at 65°C for 20min. |
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| Methylation Effect | |
| DAM, DCM, EcoKl, EcoBl | Never overlaps - no effect |
| CpG | May overlap - no effect |
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| Number of Recognition Sites in DNA | |
| Lambda | 1 |
| PhiX174, M13mp18/19,pBR322, pUC18/19, pUC57, pTZ19R/U | 0 |
| |
| Digestion of Agarose-embedded DNA | |
| A minimum of 10 units of the Bpl I is required for complete digestion of 1ug of agarose-embedded lambda DNA in 16 hours. |
|
| Overdigestion Assay | |
| No detectable change in the specific fragmentation pattern is observed after 160-fold overdigestion (10u/ug lambda DNA x 16 hours) with BplI. |
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| Ligation/Recutting Assay | |
| After 10-fold overdigestion (0.6u/ug DNA x 17 hours) with BplI, more than 70% of the DNA fragments can be ligated at a 5'-termini concentration of 0.03µM. None of these can be recut due to the methylation at the recognition sequence by Bpll. |
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| Labeled Oligonucleotide (LO) Assay | |
| No detectable degradation of a single-stranded and double-stranded labeled oligonucleotide was observed after incubation with 10 units of Bpl I for 4 hours. |
|
| Blue/White Cloning Assay | |
| The mix of pUC57/HindIII, pUC57/PstI and pUC57/Eco32I digests was incubated with 10 units of Bpl I for 16 hours. After religation and transformation the background level of white colonies was <1%. |
