Dictyostelium discoideum is a eukaryotic microbe feeding on soil bacteria. A first step towards describing the genetic structure of populations of this species was made by examining multiple isolates from a single locale. The isolates were grown clonally and their RFLP patterns compared, using a probe specific for a family of tRNA genes. Thirty-nine types were distinguished in 54 isolates. To determine if genetic exchange occurs among members of the population, an analysis of linkage disequilibrium was performed on the RFLP data. Little disequilibrium was found, implying gene flow in the population. In conflict with this result is the finding that no recombinant progeny were recovered from many attempted crosses between pairs of isolates. The tentative conclusion is that genetic exchange does not in fact occur, and that the observed shuffling of RFLP bands is caused by insertion and excision of transposons known to be associated with the tRNA genes of Dictyostelium.