Salt Active Nuclease (SAN)
Main advantages with Salt Active Nuclease
- Non specific endonuclease
- Optimum activity at high salt concentration (0.5 M NaCI)
- Active at low temperatures (20% at 6ºC)
- Broad pH range
- Temperature stable
Properties
Unit definition: One Unit is defined as an increase in absorbance at 260 nm of 0.001 per minute at 37ºC, using 50 μg/ml calf thymus DNA (D-1501, Sigma) in a buffer consisting of 25 mM Tris-HCl, pH 8.5 (25ºC), 5 mM MgCl2, 500 mM NaCl.
Specific activity: 7.6 x 106 Units/mg.
pH optimum: pH 9.
Salt optimum: 0.5 M NaCl.
Storage buffer: 25 mM Tris-HCl pH 7.5, 5 mM MgCl2, 0.5 M NaCl, 0.01% Triton, 50% (v/v) glycerol.
Reaction buffer: 25 mM Tris-HCl, pH 8.5 (25ºC), 5 mM MgCl2, 500 mM NaCl, 50 μg/ml calf thymus DNA.
Salt active Nuclease has optimum activity at high salt and pH
Relative activity of SAN at different pH and salt.
Salt active Nuclease is active in various buffers
Relative SAN activity in presence of various common buffer components in SAN enzyme assay. One hundred percent activity is set at standard assay conditions (25 mM Tris-HCl, pH 8.5, 5 mM MgCl2, 0.5 M NaCl).
SAN efficiently removes DNA from E. coli cell lysates
Removal of DNA from E. coli lysates. SAN (μg) was added to 0.1 ml E. coli cell lysate containing 75 μg/ml DNA. (Lysis buffer: 50 mM Tris-HCl, pH 8, 0,25, or 0,5 M NaCl, 0.1 mg/ml lysozyme, 10 mM MgCl2). Reactions were incubated at 2°C or 37°C for 30 minutes, followed by addition of EDTA to terminate the reactions and agarose gel electrophoresis analysis.