Mutation scanning of the RET protooncogene using high-resolution melting analysis.
Clin Chem
; 52(1): 138-41, 2006 Jan.
Article
em En
| MEDLINE
| ID: mdl-16391329
ABSTRACT
BACKGROUND:
Single-base pair missense mutations in exons 10, 11, 13, 14, 15, and 16 of the RET protooncogene are associated with the autosomal dominant multiple endocrine neoplasia type 2 (MEN2) syndromes MEN2A, MEN2B, and familial medullary thyroid carcinoma. The current widely used approach for RET mutation detection is sequencing of the exons.METHODS:
Because RET mutations are rare and the majority are heterozygous mutations, we investigated RET mutation detection by high-resolution amplicon melting analysis. This mutation scanning technique uses a saturating double-stranded nucleic acid binding dye, LCGreen, and the high-resolution melter, HR-1, to detect heterozygous and homozygous sequence variations. Mutant genotypes are distinguished from the wild-type genotype by an altered amplicon melting curve shape or position.RESULTS:
Samples of 26 unique RET mutations, 4 nonpathogenic polymorphisms, or the wild-type genotype were available for this study. The developed RET mutation-scanning assay differentiated RET sequence variations from the wild-type genotype by altered derivative melting curve shape or position. A blinded study of 80 samples (derived from the 35 mutant, polymorphism, or wild-type samples) demonstrated that 100% of RET sequence variations were differentiated from wild-type samples. For exons 11 and 13, the nonpathogenic polymorphisms could be distinguished from the pathogenic RET mutations. Some RET mutations could be directly genotyped by the mutation scanning assay because of unique derivative melting curve shapes.CONCLUSION:
RET high-resolution amplicon melting analysis is a sensitive, closed-tube assay that can detect RET protooncogene sequence variations.
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Base de dados:
MEDLINE
Assunto principal:
Proteínas Proto-Oncogênicas c-ret
Limite:
Humans
Idioma:
En
Revista:
Clin Chem
Assunto da revista:
QUIMICA CLINICA
Ano de publicação:
2006
Tipo de documento:
Article
País de afiliação:
Estados Unidos