Detection and Separation of Rhizobium and Bradyrhizobium Nod Metabolites Using Thin-Layer Chromatography. Herman P. Spaink. Department of Plant Molecular Biology, Leiden University, Nonnensteeg 3, 2311 VJ Leiden, The Netherlands. Annet Aarts(1), Gary Stacey(2), Guido V. Bloemberg(1), Ben J. J. Lugtenberg(1), and Eugene P. Kennedy(3). (1)Department of Plant Molecular Biology, Leiden University, Nonnensteeg 3, 2311 VJ Leiden, The Netherlands; (2)Center for Legume Research, Department of Microbiology and Graduate Program of Ecology, University of Tennessee, Knoxville 37996-0845 U.S.A.; (3)Department of Biological Chemistry and Molecular Pharmacology, Harvard University Medical School, Boston, MA 02115 U.S.A. MPMI 5:72-80. Accepted 7 October 1991. This article is in the public domain and not copyrightable. It may be freely reprinted with customary crediting of the source. The American Phytopathological Society, 1992.

Using radioactive acetate as a precursor, it was shown that the common nodABC genes of Rhizobium and Bradyrhizobium strains are involved in the production of one or more metabolites that are excreted into the growth medium. A rapid thin-layer chromatography (TLC) system has been developed to separate these so-called Nod metabolites that can then be visualized by autoradiography. Different patterns of Nod metabolites were observed in the tested strains of the cross-inoculation groups of R. leguminosarum bv. viceae, R. l. bv. trifolii, R. meliloti, and B. japonicum. Only Nod metabolites of R. meliloti became labeled when radioactive sulphate was present in the medium. The role of the other nodulation genes of R. l. bv. viceae in the production of the detected Nod metabolites was tested in further detail. In addition to the common nodABC genes, the nodFE and nodL genes are involved in the production of Nod metabolites. In contrast, the chromosomal background did not influence the number of detected Nod metabolites or their mobilities on TLC plates. Nod metabolites could also be produced and excreted in Escherichia coli cells in which the appropriate nodulation genes were expressed.