When using a buffer other than the optimal (supplied) NEBuffer, it may be necessary to add more enzyme to achieve complete digestion.
Heat Inactivation: 65°C for 20 minutes
Survival in a Reaction: Minimum units to digest 1 µg of substrate DNA in 16 hours: 0.50 unit(s)
Reaction & Storage Conditions Reaction Conditions: 1X NEBuffer 4 Supplemented with 20 μM S-adenosylmethionine (SAM) 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: 5,000 units/ml
Unit Assay Substrate: λ DNA
Storage Conditions: 10 mM Tris-HCl 100 mM NaCl 1 mM Dithiothreitol 0.1 mM EDTA 200 µg/ml BSA 50% Glycerol
pH 7.5 @ 25°C
Storage Temperature: -20°C
Diluent Compatibility: Diluent A
Notes General notes:
S-adenosylmethionine or SAM is supplied as a 32 mM solution in 0.005 M sulfuric acid and 10% ethanol. Under these conditions SAM is stable for up to 6 months when stored at -20°C.
The cleavage position may shift one base depending on the DNA sequence context between the recognition site and point of cleavage.
Quality Control for Current Lot Quality control values for a specific lot can be found on the datacard which accompanies each product.
16-Hour Incubation:
A 50 μl reaction containing 1 μg of λDNA
and 10 units of CspCI 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 50 units of CspCI 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.
