Genetic studies were undertaken on 14 pleiotropic negative sporulation mutants. These mutants (spoA) which are blocked early in the sporulation process were found to map near the terminus of the Bacillus subtilis chromosome in a region enriched in genes involved in spore formation. Two- and three-factor crosses by transduction and transformation led to the conclusion that the pleiotropic spoA mutations formed a linked cluster. The genetic distance across the cluster calculated from transformation data was compatible with the mutant sites defining a single gene. Suppressor studies revealed that either a nonsense or missense mutation in the spoA locus generated a pleiotropic negative phenotype. It was concluded that the locus codes for a protein, and the absence of this protein is responsible for the pleiotropic phenotype.