There are some restriction enzymes which cleave partially different base sequence than recognition sequence as a result of reduced specificity when used in a large excess to substrate DNA. This phenomenon is called Star Activity.
Conditions that Contribute to Star Activity
1. High glycerol concentration [>5% v/v]
2. High units to µg of DNA ratio [Varies with each enzyme, usually >100 units/µg]
3. Low ionic strength [<25 mM]
4. High pH [>pH 8.0]
5. Presence of organic solvents [DMSO, ethanol (9), ethylene glycol, dimethylacetamide, dimethylformamide, sulphalane (12)]
6. Substitution of Mg++ with other divalent cations [Mn++, Cu++, Co++, Zn++]
The relative significance of each of these altered conditions is dependent on the enzyme in question. For example, EcoR I is much more sensitive to elevated glycerol concentrations than is Pst I, which is more sensitive to elevated pH (2).