| 5'-Pu^A A T T Py-3' |
|
| 3'-Py T T A A^Pu-5' |
|
| Catalog # | X0985 |
| Source | Xanthomonas ampelina Slo 51–021 |
| Concentration | 10u/ul |
| Form | Supplied as a liquid (Storage Buffer) in 10mM Tris-HCl, pH 7.4, 100mM KCl, 1mM DTT, 1mM EDTA, 0.2mg/ml BSA and 50% glycerol. |
| Restriction Enzyme Buffer A, 10X (for 100% digestion) R1625 | 33mM Tris-acetate, pH 7.9, 10mM magnesium acetate, 66mM potassium acetate and 0.1mg/ml BSA. Incubate at 37°C. |
| Unit Definition | One unit is defined as the amount of XapI required to digest 1ug of lambda DNA in 1 hour at 37°C in 50ul of assay buffer.. |
| Storage and Stability | May be stored at 4°C for short-term only. For long-term storage, store at -20°C. Aliquots are stable for at least 6 months at -20°C. For maximum recovery of product, centrifuge the original vial after thawing and prior to removing the cap. Further dilutions can be made in assay buffer. |
| Overdigestion Assay | |
| No detectable change in the specific fragmentation pattern is observed after 15-fold overdigestion with XapI(15u/ug lambda DNA x 1 hours) . |
|
| Ligation/Recutting Assay | |
| After 10-fold overdigestion (5u/ug DNA x 2 hours) with HpaII, more than 95% of the DNA fragments can be ligated at a 5'-termini concentration of 0.2uM. More than 95% of these can be recut. |
|
| Labeled Oligonucleotide (LO) Assay | No detectable degradation of single-stranded and double-stranded labeled oligonucleotide was observed after incubation with 10 units of XapI for 4 hours. |
|
| Blue/White Cloning Assay | |
| pUC57 was digested at a unique site with 10 units of XapI for 1 hours. After religation and transformation 0.2 % of white colonies were detected. |
| |
| Star Activity | An excess of XapI (20u/ug DNA x 1 hour) may result in star activity. |
|
| Stability during Prolonged Incubation | A minimum of 0.1 units of XapI is required for complete digestion of 1ug of lambda DNA in 16 hours at 37°C. |
|
| Digestion of Agarose-embedded DNA | A minimum of 5 units of XapI is required for digestion of 1ug of agarose-embedded lambda DNA in 16 hours. |
|
| Compatible Ends | |
| EcoRI, MunI, TasI |
|
| Number of Recognition Sites in DNA | |
| Lambda | 58PhiX174: 5 |
| pBR322 | 1 |
| pUC57 | 1 |
| pUC18/19 | 1 |
| pTZ19R/U | 3 |
| M13mp18/19 | 11 |
|
| Protocol for Digestion | |
| Add | 44ul nudlease free dH2O |
| 5ul Restriction Enzyme Buffer A, 10X |
| 1ul DNA (0.5-1ug/ul) |
| 0.5-2ul XapI |
| Mix gently and spin down for a few seconds. |
| Incubate at 3ºC for 1-2 hours. |
|
| Protocol for Digestion of PCR products Directly after Amplification | |
| Add | 10ul (~1ug DNA) |
| 16ul nuclease free dH2O |
| 2ul Restriction Enzyme Buffer A, 10X |
| 1-2ul XapI |
| Mix gently and spin down for a few seconds. |
| Incubate at 3ºC for 1-16 hours. |
|
| Thermal Inactivation | |
| XapI is inactivated by incubation at 65°C for 20min. |
