The polymorphic DNA probe D3S3 was assigned to 3pl4 by molecular hybridization using a human chromosome 3/hamster somatic cell hybrid deletion panel. This is the first regional assignment of a polymorphic probe to the short arm of chromosome 3. This probe appears to be proximal to the chromosome 3 fragile site and, therefore, may prove useful in characterizing the 3p chromosomal aberrations that occur in various malignant diseases.