| 5'-A^G A T C T-3' |
|
| 3'-T C T A G^A-5' |
|
| Catalog # | B1087-05 |
| Enzyme Properties | Methylation Effects: |
| Dam: completely overlaps-no effect. |
| Dcm/CpG/EcoKl: never overlaps-no effect. |
| EcoBl: may overlap-cleavage impaired. |
| Stability during Prolonged Incubation | A minimum of 0.1 units of enzyme is required for complete digestion of 1ug of lambda DNA in 16 hours at 37°C. |
| Thermal Inactivation | Enzyme is not inactivated by incubation at 80°C for 20 minutes. |
| Digestion of Agarose-embedded DNA | A minimum of 5 units of enzyme is required for digestion of 1ug of agarose-embedded lambda DNA in 16 hours. |
| Compatible Ends | BamHI, BclI, Bsp143I, MboI, PsuI |
| Number of Recognition Sites in DNA | Lambda: 6 |
| PhiX174: 0 |
| M13mp18/19: 1 |
| pBR322: 0 |
| pUC18/19: 0 |
| pUC57: 0 |
| pTZ19R/U: 0 |
| |
| Concentration | 10u/ul |
|
| Source | Bacillus globigii |
|
| Form; |
| Supplied as aliquid in 50mM Tris-HCl pH 7.5, 10mM MgCl |
|
| Diluent Buffer | |
| For short term storage (3-4 weeks, dilute with Diluent Buffer (10mM Tris-HCl, pH 7.4,,100mM potassium chloride, 1mM EDTA, 1mM DTT, 0.2mg/ml BSA, 50% glycerol). |
|
| Storage Buffer | |
| For long term storage, use Storage Buffer (10mM Tris-HCl, pH 8.2, 200mM potassium chloride, 1mM DTT, 0.1mM EDTA, 0.5mg/ml BSA, 50% glycerol). |
|
| Quality Control | |
| Overdigestion Assay | |
| No detectable change in the specific fragmentation pattern is observed after 160-fold overdigestion (10u/ug lambda DNA x 16 hours) with BglII. |
|
| Ligation/Recutting Assay | |
| After 50-fold overdigestion (3u/ug DNA x 17 hours) with BglII, 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 10units of BglII for 4 hours. |
|
| Blue/White Cloning Assay | |
| The mix of pUC57/HindIII, pUC57/PstI and pUC57/Eco32I digests was incubated with 10units of BglII for 16 hours. After religation and transformation <1% of white colonies were detected. |
|
| Protocol for Digestion | |
| Add | |
| Nuclease free water | 16ul |
| R1625-03 | 2ul |
| DNA (0.5-1ug/ml) | 1ul |
| B1087-05 | 0.5-2ul |
| Mix gently and spin down for a few seconds. Incubate at 37ºC for 1-16 hours. |
|
| Protocol for Digestion Directly after Amplification | |
| Add | |
| PCR Reaction Mixture | 10ul (~0.1-0.5ug of DNA) |
| Nuclease free water | 18ul |
| R1625-03 | 2ul |
| B1087-05 | 1-2ul |
| Mix gently and spin down for a few seconds. |
| Incubate at 37ºC for 1-16 hours. |
