title: Application of dHPLC for mutation detection of the fibrillin-1 gene for the diagnosis of Marfan syndrome in a National Health Service Laboratory.
abstract: Marfan syndrome (MFS) is an autosomal dominant connective tissue disorder caused by mutations in the fibrillin-1 gene FBN1. Mutation detection of this 65-exon gene presents a particular challenge for the diagnostic service in cost, time constraints, and the need to maintain a stringently optimized assay procedure. Using denaturing high-performance liquid chromatography (dHPLC), we have designed a procedure for rapid mutation scanning, redesigning 50% of published primer sets, screening by Ensembl to avoid inclusion of polymorphic variations and employing a limited set of PCR conditions to allow for a high-throughput 96-well format. We have screened 262 unrelated patients with MFS or Marfan-like phenotypes and detected 103 (39.3%) mutations including 93 different mutations, 72 of which are novel. The mutations include 55 missense (53.4%) 19 splice site (18.5%), 17 frameshift (16.5%), 11 nonsense (10.7%) and 1 in-frame deletion/insertion.
authors.affiliation: Wessex Regional Genetics Laboratory, Salisbury Hospital NHS Trust, Salisbury, Wiltshire, SP2 8BJ, UK.
authors.forename: Rachel
authors.initials: R
authors.lastname: Howarth
authors.forename: Catharina
authors.initials: C
authors.lastname: Yearwood
authors.forename: John F
authors.initials: JF
authors.lastname: Harvey
mesh_terms: D019943:Amino Acid Substitution; D002851:Chromatography, High Pressure Liquid / Q000379:methods; D004247:DNA / Q000235:genetics / Q000302:isolation & purification; D017931:DNA Primers; D004739:England; D005091:Exons; D000071838:Fibrillin-1; D000071837:Fibrillins; D006801:Humans; D008382:Marfan Syndrome / Q000175:diagnosis* / Q000235:genetics*; D008840:Microfilament Proteins / Q000235:genetics*; D009154:Mutation; D016133:Polymerase Chain Reaction; D017384:Sequence Deletion; D013222:State Medicine; D054392:Adipokines
chemical_list: D004247:DNA; D017931:DNA Primers; D000071838:Fibrillin-1; D000071837:Fibrillins; D008840:Microfilament Proteins; C000605722:FBN1 protein, human; D054392:Adipokines