| 5-A G T A C T-3 3-T C A T G A-5 Concentration: 10u/ul Source: Streptomyces caespitosus Supplied with: R1625-70: Restriction Enzyme Buffer for ScaI, SdaI, 10X 37mM Tris-acetate (pH 7.0), 15mM magnesium acetate, 150mM 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. For longer periods-the Storage Buffer should be used. Storage Buffer: 10mM Tris-HCl (pH 7.5 at 25C), 50mM KCl, 1mM DTT, 0.1mM EDTA, 0.5mg/ml BSA and 50% glycerol. Overdigestion Assay: No detectable change in the specific fragmentation pattern is observed after 5-fold overdigestion (5u/ug lambda DNA x 1 hour) with ScaI (see Star Activity). Ligation/Recutting Assay: After 5-fold overdigestion (5u/ug DNA x 1 hour) with ScaI, more than 80% of the DNA fragments can be ligated at a 5-termini concentration of 0.05uM. 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 (7.5u/ug DNA x 1 hour) may result in star activity. Stability during Prolonged Incubation: A minimum of 0.3 units of enzyme is required for complete digestion of 1ug of lambda DNA in 16 hours at 37C. Thermal Inactivation: Only small amounts of enzyme (up to 10 units) can be inactivated by incubation at 80C for 20min. Digestion of Agarose-embedded DNA: A minimum of 20 units of enzyme is required for digestion of 1ug of agarose-embedded lambda DNA in 16 hours. Number of Recognition Sites in DNA: Lambda: 5 PhiX174: 0 M13mp18/19: 0 pBR322: 1 pUC18/19: 1 pUC57: 1 pTZ19R/U: 1 pBluescriptIIKS(-/+): 1 pBluescriptIISK(-/+): 1 pACYC177: 1 pACYC184: 1 5 0 0 1 1 1 1 1 1 1 1 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. NR: buffer is not recommended, since star activity appears at 5-fold overdigestion (5 units x 1 hour). |