Binding of DNA on humic acids: Effect on transformation of Bacillus subtilis and resistance to DNase

Abstract

Equilibrium adsorption and binding of DNA from Bacillus subtilis on humic acids (HA) extracted from a forest soil, the capacity of the bound DNA to transform B. subtilis, and the resistance of the bound DNA to degradation by DNase I are reported. Adsorption of DNA on the HA was maximal after 2 and 4 h at pH 3.0 and 4.0, respectively. The adsorption of a constant amount of DNA (50 μg) on increasing concentrations of the HA reached a plateau with 2 and 3 mg of HA at pH 3.0 and 4.0, respectively. No plateau was observed when increasing amounts of DNA where adsorbed on a constant amount of the HA (2 mg). 70 to 80% of the adsorbed DNA was tightly bound after two washes with 0.1 M NaCl (pH 6.4), double distilled water (pH 5.5), or DNA buffer (pH 4.0). Bound DNA was capable of transforming auxotrophic and chloramphenicol-sensitive cells of B. subtilis, although at a lower frequency than free DNA. DNA bound on the HA was protected more against degradation by DNase I than free DNA: the concentration of DNase required to inhibit transformation by bound DNA was approximately 100 times higher than that required to inhibit transformation by comparable amounts of free DNA.