The recessive gene rhm confers chlorotic-lesion resistance to Bipolaris maydis race O, the southern corn leaf blight pathogen, in otherwise susceptible maize plants. Because of inconsistencies encountered in scoring the disease on mature plants in the field, an assay was developed to monitor the expression of this gene in maize seedlings under controlled conditions. One hundred and two related F3 families from the cross RH95rhm X B73 were inoculated with conidia of B. maydis race O, and the genotype at the rhm locus of each F2 parent was deduced from the reactions observed in the progeny seedlings. The F2 genomes were reconstituted by extracting DNA from leaf tissue pooled from 30-36 F3 progeny plants per family. The seedling disease ratings were analyzed together with the segregation scores for 14 single-copy DNA probes. Our results indicate that rhm is tightly linked to two restriction fragment length polymorphism (RFLP) marker loci (UMC85 and p144) that map to the short arm of chromosome 6. In addition, a rapid assay based on the polymerase chain reaction was used to confirm the linkage between rhm and the p144 RFLP marker locus in a second unrelated F2 population.