KpnI

Produktübersicht

• Artikelname: KpnI
• Artikelnummer: USB-K1894
• Hersteller Artikelnummer: K1894
• Alternativnummer: USB-K1894-4000
• Hersteller: US Biological
• Kategorie: Molekularbiologie

Produktbeschreibung

5-G G T A C C-3 3-C C A T G G-5 Source: Klebsiella pneumoniae OK8 Concentration: 10u/ul Unit Definition: One unit is defined as the amount of KpnI required to digest 1ug of lambda DNA-BamHI fragmants in 1 hour at 37C in 50ul of assay buffer Supplied With: R1625: Restriction Enzyme Buffer A, 10X: Supplied as a liquid in 33mM Tris-acetate, 10mM magnesium acetate, 66mM potassium acetate, 0.1mg/ml BSA. R1625-65: Restriction Enzyme Buffer for KpnI, 10X: Supplied as a liquid in 10mM Tris-HCl, pH 7.5, 10mM MgCl2, 0.02% Triton X-100 and 0.1mg/ml BSA. Incubate at 37C. Dilution Buffer: 10mM Tris-HCl, pH 7.4, 100mM KCl, 1mM EDTA, 1mM DTT, 0.2mg/ml BSA and 50% glycerol. Storage Buffer: Supplied as a liquid in10mM Tris-HCl (pH 7.5 at 25C), 50mM KCl, 1mM DTT, 0.1mM EDTA, 0.2mg/ml BSA and 50% glycerol. Thermal Inactivation: KpnI is inactivated by incubation at 80C for 20min. Overdigestion Assay: No detectable change in the specific fragmentation pattern is observed after 80-fold overdigestion (10u/ug lambda DNA x 16 hours) with KpnI. Ligation/Recleavage (L/R) Assay: The ligation and recleavage assay was replaced with LO test after validating experiments showed LO test ability to trace nuclease and phosphatase activities with sensitivity that is higher than L/R by a factor of 100. Labeled Oligonucleotide (LO) Assay: No detectable degradation of single- stranded or double-stranded labeled oligonucleotide was observed after incubation with 10 units of KpnI for 4 hours. Blue/White Cloning Assay: The B/W assay was replaced with LO test after validating experiments showed LO test ability to detect nuclease and phosphatase activities with sensitivity that equals to that of B/W test. Stability during Prolonged Incubation: A minimum of 0.2 units of KpnI is required for complete digestion of 1ug of lambda DNA in 16 hours at 37C. Number of Recognition Sites in DNA: Lambda: 2: M13mp18/19, pUC18/19, pUC57, pTZ19R/U: 1 PhiX174, pBR322: 0 Digestion of Agarose-embedded DNA: A minimum of 5 units of KpnI is required for digestion of 1ug of agarose-embedded lambda DNA in 16 hours. Methylation Effects: Dam, EcoBl, EcoKl: never overlaps - no effect Dcm, CpG: may overlap - no effect Recommended Protocol for Digestion: Add: Nuclease free water: 16ul R1625-65: 2ul DNA (0.5-1ug/ul): 1ul KpnI: 0.5-2ul Mix gently, spin down for a few seconds. Incubate at 37C for 1-16 hours. Protocol for Digestion of PCR products directly after amplification: Add: PCR Reaction Mixture: 10ul (~0.1-0.5ug of DNA) Nuclease free water: 18ul R1625-65: 2ul KpnI 1-2ul Mix gently, spin down for a few seconds. Incubate at 37C for 1-16 hours. Storage and Stability: May be stored at 4C for short-term only. For long-term storage, store at -20C. Aliquots are stable for 6 months after receipt at -20C. For maximum recovery of product, centrifuge the original vial after thawing and prior to removing the cap.