COLD SINGLE-STRAND CONFORMATIONAL VARIANTS FOR MUTATION ANALYSIS OF THE RET PROTOONCOGENE

Background. RET protooncogene mutation analysis is a routinely perform ed predictive DNA test in kindreds affected by multiple endocrine neop lasia (MEN) types 2A and 2B and familial medullary thyroid carcinoma ( FMTC) and is a valuable diagnostic tool in newly diagnosed cases of me dullary thyroid carcinoma (MTC).Methods. We tested the suitability of the recently introduced ''cold'' single-stand conformational variant ( SSCV) technique, which promises rapid simple, nonradioactive detection of sequence variants in the identification of germline and somatic RE T mutations. A total of II different mutations in exon 10 (codons 609, 611, 628, and 620) and 6 mutations in exon 11 (codon 634) were studie d.Results. Conditions were optimized so that conformational variants w ere demonstrated for all mutations examined in a single setting for ex ons 10 and 11. A novel six base pair (bp) inframe deletion between cys teines 630 and 634 was detected in a sporadic MTC. This adds to the ev idence that not only cysteine deletions and substitutions but also cha nges in the spacing between cysteine residues have a pathogenic effect . Conclusions. Our results indicate that the cold SSCV method offers t he advantages of simplicity, time savings, and nonradioactive detectio n for screening for RET sequence variants in hereditary and sporadic M TCs.