Home Protocols General Guidance for NEBExpress® Salt Active Nuclease Usage (NEB #M0764)

General Guidance for NEBExpress® Salt Active Nuclease Usage (NEB #M0764)

Optimal enzyme concentration and incubation times may vary depending on the sample preparation (pH, salt concentration, DNA and RNA concentration), the reaction temperature (typically between 4°C and 37°C), and the application.

For samples prepared in 50 mM Tris-HCl pH 8.5, 500 mM NaCl, 5 mM MgCl2, we recommend starting with:

 
Application Sample type Unit per mL of reaction Conditions Tips
Viscosity reduction Cell lysate** 10 U 10 min at 25°C
  • Increase the amount of enzyme or incubation time if:
    -   pH <8.5
    -   NaCl < or > 500 mM
    -   temperature < 25°C
  • Decrease units of enzyme or incubation time if the reaction temperature is >25°C
Nucleic acid removal* Purified sample 10 U 30 min at 37°C 
  100 U 30 min at 25°C
  1,000 U 30 min at 4°C
Soluble fraction** 10 U 2 hrs at 37°C  
  100 U
 2 hrs at 25°C
   1,000 U
 2 hrs at 4°C
Cell lysate** 40 U 2 hrs at 37°C 
  400 U
 2 hrs at 25°C
  2,500 U
 2 hrs at 4°C

*DNA not visually detectable on agarose gel
** All samples tested were from E. coli cells (1 g of wet cells resuspended in 5 mL of buffer)

Note: One unit of NEBExpress Salt Active Nuclease will digest 30 µg of calf thymus DNA in 30 minutes at 37°C in 25 mM Tris-HCl, 500 mM NaCl, 5 mM MgCl2, pH 8.5.