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FAQs for ScaI-HF™ FAQs for Restriction Endonucleases Technical Reference for Restriction Endonucleases Enzyme Finder NEBcutter NEBuffer Chart Double Digest Finder Isoschizomers DNA Sequences and Maps REBASE NEBuffer 4

Home > Products > Restriction Endonucleases > Restriction Endonucleases > ScaI-HF™ ScaI-HF™ High-Fidelity Enzymes Catalog # Size Concentration Price Qty   R3122S 1,000 units 20,000 units/ml $58.00 Prices are in US dollars and valid only for US orders. Download: MSDS PDF Recognition Site: isoschizomers | compatible ends | single letter code Description: Many restriction endonucleases are capable of cleaving sequences which are similar but not identical to their defined recognition sequence. This altered or relaxed specificity has been termed star activity. HF restriction endonucleases have been engineered to cleave with higher fidelity than the wild type enzyme, hence exhibiting less star activity. Source: An E. coli strain that carries the cloned and modified (H193A/S201F) ScaI gene from Streptomyces caespitosus (H. Takahashi) Reagents Supplied: NEBuffer 4 Enzyme Properties Activity in NEBuffers: NEBuffer 1: 100% NEBuffer 2: 100% NEBuffer 3: 10% NEBuffer 4: 100% When using a buffer other than the optimal (supplied) NEBuffer, it may be necessary to add more enzyme to achieve complete digestion. Methylation Sensitivity: dam methylation: Not sensitive dcm methylation: Not sensitive CpG methylation: Not sensitive Heat Inactivation: 65°C for 20 minutes Survival in a Reaction: Minimum units to digest 1 µg of substrate DNA in 16 hours: 0.25 unit(s) Reaction & Storage Conditions Reaction Conditions: 1X NEBuffer 4 Incubate at 37°C. 1X NEBuffer 4: 20 mM Tris-acetate 50 mM potassium acetate 10 mM Magnesium Acetate 1 mM Dithiothreitol pH 7.9 @ 25°C Unit Definition: One unit is defined as the amount of enzyme required to digest 1 μg of λ DNA in 1 hour at 37°C in a total reaction volume of 50 μl. Concentration: 20,000 units/ml Unit Assay Substrate: λ DNA Storage Conditions: 10 mM Tris-HCl 200 mM NaCl 1 mM Dithiothreitol 0.1 mM EDTA 200 µg/ml BSA 50% Glycerol pH 7.4 @ 25°C Storage Temperature: -20°C Diluent Compatibility: Diluent B Notes Usage notes: ScaI-HF™ has the same specificity as ScaI (NEB #R0122), but it has been engineered for reduced star activity. FAQs What is the specific activity of ScaI-HF™? What does HF refer to following the name of the restriction enzyme? Why does the HF version of the enzyme have a different recommended buffer than the wild type enzyme? Can the change in buffer preference of the HF enzyme be advantageous? Will the HF enzyme produce elevated star activity when used in a buffer other than the one recommended? Is there any difference in the methylation sensitivity between ScaI-HF and ScaI? What is the difference between ScaI-HF and ScaI? When should I choose the HF version of the enzyme? When is star activity a concern? Is there a difference in cutting close to the ends between ScaI-HF and ScaI? What does it mean to be Time-Saver qualified? Quality Control for Current Lot Quality control values for a specific lot can be found on the datacard which accompanies each product. Ligation and Re-cutting: After a 2-fold overdigestion with ScaI-HF™, > 95% of the DNA fragments can be ligated with T4 DNA Ligase (at a 5' termini concentration of 1-2 μM) at 16ºC. Of these ligated fragments, > 95% can be recut with ScaI-HF™. 16-Hour Incubation: A 50 μl reaction containing 1 μg of DNA and 60 units of ScaI-HF™ incubated for 16 hours at 37ºC resulted in a DNA pattern free of detectable nuclease degradation as determined by agarose gel electrophoresis. Exonuclease Activity: Incubation of a 50 μl reaction containing 200 units of ScaI-HF™ with 1 μg of a mixture of single and double-stranded [3H] E. coli DNA (205 cpm/μg) for 4 hours at 37ºC released < 0.1% of the total radioactivity. Endonuclease Activity: Incubation of a 50 μl reaction containing 100 units of ScaI-HF™ with 1 μg of ΦX174 RF I DNA for 4 hours at 37ºC resulted in < 50% conversion to RFII as determined by agarose gel electrophoresis. Reagents Sold Separately NEBuffer 4 Legal Patents: New England Biolabs, Inc.: U.S. Patent No. 5,721,126