POLYMERASE CHAIN REACTION-BASED TECHNIQUE FOR THE SELECTIVE ENRICHMENT AND ANALYSIS OF MOSAIC ARG(201) MUTATIONS IN G-ALPHA(S) FROM PATIENTS WITH FIBROUS DYSPLASIA OF BONE

Mutations in the arg(201) codon of the alpha(s) G protein subunit have been associated with a variety of disorders, but analysis of such mut ations has been complicated by their mosaic presentation, To overcome the problems associated with the analysis of genomic mutations that ma y be present in low and variable yield throughout the body, a polymera se chain reaction (PCR)-based technique has been developed that allows the selective amplification of products from the mutant allele, This technique uses site-directed mutagenesis to generate a PCR product fro m the normal allele that is susceptible to restriction endonuclease di gestion, whereas that from the mutant allele is resistant to digestion , Consecutive and repeated cycles of amplification and digestion allow selective enrichment of the product from the mutant allele, The techn ique has been applied to the analysis of patients with fibrous dysplas ia of bone, where the consequence of Ga, mutations may vary from monos totic to polyostotic lesions, and has been performed with DNA isolated from either bone biopsy specimens or peripheral blood leukocytes, In addition to the previously described arg --> his and arg --> cys subst itutions, the analyses have detected a novel arg --> ser substitution in one of the patients, This patient presented with a panostotic disea se and may represent a unique subgroup of fibrous dysplasia. (C) 1997 by Elsevier Science Inc. All rights reserved.