Improvement in the RFLP procedure for studying the diversity of nifH genes in communities of nitrogen fixers in soil.

Several specific primers for the nifH gene were tested with different pure telluric N2-fixing strains. A PolF/PolR primer set provided successful amplification of 19 representative N2-fixing strains. Three restriction enzymes, HaeIII, NdeII and MnlI, chosen for restriction fragment length polymorphism (RFLP) analyses, were the most discriminating for the study of nifH gene diversity as they resulted in differences between strains at the species level. Amplification by selected primers and RFLP were applied to assess the genetic diversity of the nifH gene pool in soil. Pair soils, one under cultivation, the second under permanent pasture, were found to harbor a contrasting diversity of nifH genes. Pure strain profiles could not be recognized in the nifH soil patterns. Using the simple procedure described, it was shown that the structure of nitrogen fixers in soil was influenced by soil functioning.