Osteogenesis imperfecta (OI) is a heterogeneous group of inherited diseases of connective tissue manifested primarily by excessive fracturing of bone but associated with other abnormalities such as blue sclera, thin skin, herniae, ligamentous laxity, reduced stature and hearing loss. We report here molecular studies on a patient with the mild, dominantly inherited, variety of OI (OI type I) previously shown (Nicholls et al., 1984) to be heterozygous for an abnormal alpha 1(I) chain of type I collagen which contained cysteine near the carboxyl terminus (Steinmann et al., 1986). The cognate alpha 1(I) mRNA region was selected for generation of cDNAs which were subsequently amplified by the polymerase chain reaction (PCR), cloned and sequenced. Two sequences were obtained, one of which corresponded to the normal allele, and the other of which harbored a G to T transversion and resulted in a cysteine for glycine substitution. This is the first single amino acid substitution found in type I OI. Surprisingly, the mutation occurs just outside the triple-helical region of the alpha 1(I) chain, a result that accounts for the strikingly different phenotypic and molecular consequences of this mutation as compared with similar cysteine for glycine substitutions within the same region. The PCR appears to be a useful approach for elucidation of structural mutations in collagen chains.