In the search for the prevalence and distribution pattern of Gs-alpha gene mutations in differentiated thyroid tumors we examined 66 tumor tissue samples for the presence of mutations at "hot-spot" codons 201 and 227 using methods based on the polymerase chain reaction, subcloning and sequencing. The prevailing type of single-base substitution at codon 201 (71.4%) corresponded to the replacement of the wild-type sequence CGT (Arg) with TGT (Cys). The fragments of the Gs-alpha gene, including codon 201 or 227 from five follicular carcinomas and one follicular adenoma, were subcloned in Escherichia coli and it was found that the proportion of alleles with mutated codon 201 varied from 3.2% to 43%. Sequencing of the corresponding region has confirmed preliminary data indicating that the single-base changes CGT (Arg) to TGT (Cys) or CGT to CAT (His) occurred. There was only a weak correlation between the prevalence of cells bearing a mutation in the Gs-alpha gene and the level of Gs-alpha protein expression in the corresponding thyroid tumors.