Sequence: 5-G G N C C-3 3-C C N G G-5 Restriction Enzyme Buffer A, 10X (R1625): Supplied to simplify buffer selection for double digests. Enzyme Properties: Number of Recognition Sites in DNA: Lambda: 74 PhiX174: 2 pBR322: 15 pUC57: 8 pUC18/19: 6 pTZ19R/U: 6 M13mp18/19: 4 Methylation Effects: Cfr13I does not cut GGNCm5C. Blocked by overlapping Dcm and CG methylation. Stability during Prolonged Incubation: A minimum of 0.3units 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. Compatible Ends: CpoI, Eco47I, Psp5II, SanDI: G G(A/T)CC 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 Cfr13I. Ligation/Recutting Assay: After 50-fold overdigestion (3u/ug DNA x 17 hours) with Cfr13I, more than 95% of the DNA fragments can be ligated at a 5-termini concentration of 0.3uM. 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 restriction endonuclease for 4 hours. Storage and Stability: May be stored at 4C for short-term only. For long-term storage, store at -20C. Aliquots are stable for at least 6-12months at -20C. 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.
Form:
Supplied as a liquid in 10mM potassium phosphate, pH 7.5 at 25C, 100mM potassium chloride, 1mM EDTA, 7mM 2-mercaptoethanol, 0.2mg/ml BSA, 50% glycerol.
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