Charcot-Marie-Tooth disease type 1 A (CMTi A) was localized by genetic mapping to a 3 CM interval on human chromosome 17~. DNA markers within this interval revealed a duplication that is completely linked and associated with CMTl A. The duplication was demonstrated in affected individuals by the presence of three alleles at a highly polymorphic locus, by dosage differences at RFLP alleles, and by two-color fluorescence in situ hybridization. Pulsed-field gel electrophoresis of genomic DNA from patients of different ethnic origins showed a novel Sacll fragment of 500 kb associated with CMTlA. A severely affected CMTIA offspring from a mating between two affected individuals was demonstrated to have this duplication present on each chromosome 17. We have demonstrated that failure to recognize the molecular duplication can lead to misinterpretation of marker genotypes for affected individuals, identification of false recombinant!% and incorrect localization of the disease locus.