title: Efficient testing of the RET gene by DHPLC analysis for MEN 2 syndrome in a cohort of patients.
abstract: BACKGROUND
  Multiple Endocrine Neoplasia type 2 (MEN 2) is an autosomal dominant inherited syndrome characterized by a strong predisposition for developing endocrine tumors. MEN 2 is caused by germline mutations in the ret proto-oncogene. We investigated the feasibility of using the DHPLC technique in mutation detection of the ret gene in members of MTC families. We compared DHPLC analysis with direct sequencing with regard to sensitivity, reliability, cost and time.
  
  
  METHODS
  Exons 10 and 11 were amplified with PCR from forty-three samples in seventeen unrelated Greek families and were analyzed for mutations by DHPLC and DNA sequencing.
  
  
  RESULTS
  Eight PCR amplicons showed a distinct non-wild-type DHPLC profile. Sequence analysis confirmed different nucleotide variations: six of them were localized in exon 10 and two in exon 11. Mutations were detected in five out of seventeen families tested (29%).
  
  
  CONCLUSIONS
  None of the alterations detected by direct sequencing was missed by DHPLC. We conclude that DHPLC is a fast, sensitive, cost-efficient and reliable method for the scanning of ret germline mutations.
authors.affiliation: Department of Endocrinology and Endocrine Oncology, Theagenio Hospital, Thessaloniki, Athens, Greece.
authors.forename: Kalliopi
authors.initials: K
authors.lastname: Pazaitou-Panayiotou
authors.forename: Athina
authors.initials: A
authors.lastname: Kaprara
authors.forename: Leda
authors.initials: L
authors.lastname: Sarika
authors.forename: Thanasis
authors.initials: T
authors.lastname: Zovoilis
authors.forename: Ioulia
authors.initials: I
authors.lastname: Belogianni
authors.forename: Iraklis
authors.initials: I
authors.lastname: Vainas
authors.forename: Georgios
authors.initials: G
authors.lastname: Nasioulas
mesh_terms: D002851:Chromatography, High Pressure Liquid / Q000379:methods; D015331:Cohort Studies; D005091:Exons; D005260:Female; D018095:Germ-Line Mutation; D006801:Humans; D008297:Male; D018813:Multiple Endocrine Neoplasia Type 2a / Q000235:genetics*; D018814:Multiple Endocrine Neoplasia Type 2b / Q000235:genetics*; D015513:Oncogene Proteins / Q000235:genetics*; D010375:Pedigree; D016133:Polymerase Chain Reaction; D000090063:Proto-Oncogene Mas; D051096:Proto-Oncogene Proteins c-ret; D020794:Receptor Protein-Tyrosine Kinases / Q000235:genetics*; D017422:Sequence Analysis, DNA
chemical_list: C000715470:MAS1 protein, human; D015513:Oncogene Proteins; D000090063:Proto-Oncogene Mas; D051096:Proto-Oncogene Proteins c-ret; C099282:RET protein, human; D020794:Receptor Protein-Tyrosine Kinases