| 5-G A G T C N N N N N -3 3-C T C A G N N N N N -5 Concentration: 2u/ul Source: Staphylococcus cohnii Lki 19-320 Buffer: 33mM Tris-acetate (pH 7.9), 10mM magnesium acetate, 66mM potassium acetate and 0.1mg/ml BSA. Incubate at 37C. Diluent Buffer: 10mM Tris-HCl (pH 7.4 at 25C), 100mM KCl, 1mM EDTA, 1mM DTT, 0.2mg/ml BSA and 50% glycerol, or Storage Buffer. Storage Buffer: 10mM Tris-HCl (pH 7.5 at 25C), 100mM KCl, 1mM DTT, 0.1mM EDTA, 0.2mg/ml BSA and 50% glycerol. Overdigestion Assay: No detectable change in the specific fragmentation pattern is observed after 48-fold overdigestion (3u/ug lambda DNA x 16 hours) with SchI (see Star Activity). Ligation/Recutting Assay: After 17-fold overdigestion (1u/ug DNA x 17 hours) with SchI, approximately 90% of the DNA fragments can be ligated at a 5-termini concentration of 0.6uM. 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 10 units of restriction endonuclease for 4 hours. Star Activity: A large excess of enzyme (4u/ug DNA x 16 hours) may result in star activity. Stability during Prolonged Incubation: A minimum of 0.2 units of enzyme is required for complete digestion of 1ug of lambda DNA in 16 hours at 37C. Thermal Inactivation: Enzyme is inactivated by incubation at 65C for 20min. Number of Recognition Sites in DNA: Lambda: 61 PhiX174: 10 M13mp18/19: 8 pBR322: 4 pUC18/19: 4 pUC57: 3 pTZ19R/U: 7 pBluescriptIIKS(-/+): 6 pBluescriptIISK(-/+): 6 pACYC177: 6 pACYC184: 4 Unit Definition: One unit is defined as the amount of enzyme required to digest 1ug of lambda DNA in 1 hour at 37C in 50ul of assay buffer. |