title: Scanning mutations of the 5'UTR regulatory sequence of L-ferritin by denaturing high-performance liquid chromatography: identification of new mutations.
abstract: Hereditary hyperferritinaemia cataract syndrome is an autosomal dominant disorder caused by heterogeneous mutations of the iron regulatory element (IRE) in the ferritin l-chain mRNA. The mutations are rare and fast DNA scanning would facilitate diagnosis. The aim of the study was to compare the analytical performances of two fast DNA scanning techniques: denaturing high-performance liquid chromatography (DHPLC) and double-gradient denaturing gradient gel electrophoresis (DG-DGGE). We analysed the sequence encoding the 5' untranslated flanking region of ferritin l-chain mRNA, which includes an IRE stem loop structure. The two systems unambiguously identified all the 12 accessible mutations in a single run, including the difficult C-G transversions. DHPLC and DG-DGGE identified seven abnormal patterns in DNA samples from 47 subjects with unexplained hyperferritinaemia; all had mutations in the IRE sequence, including two not reported before: C36G and A37G. The scanning of 250 DNA samples from subjects genotyped for HFE led to the identification of four new mutations, all outside the IRE structure: C10T, C16T, C90T and del-T156. We conclude that DHPLC, similar to DG-DGGE, detects all the mutations in the l-ferritin 5'UTR sequence in a single run, and that various mutations occur outside the IRE structure.
authors.affiliation: Unit of Genetics and Molecular Diagnostics, Istituti di Ricovero e Cura a Carattere Scientifico (IRCCS), H. San Raffaele, Milan, Italy.
authors.forename: Laura
authors.initials: L
authors.lastname: Cremonesi
authors.forename: Rita
authors.initials: R
authors.lastname: Paroni
authors.forename: Barbara
authors.initials: B
authors.lastname: Foglieni
authors.forename: Silvia
authors.initials: S
authors.lastname: Galbiati
authors.forename: Isabella
authors.initials: I
authors.lastname: Fermo
authors.forename: Nadia
authors.initials: N
authors.lastname: Soriani
authors.forename: Silvana
authors.initials: S
authors.lastname: Belloli
authors.forename: Giuseppina
authors.initials: G
authors.lastname: Ruggeri
authors.forename: Giorgio
authors.initials: G
authors.lastname: Biasiotto
authors.forename: Mario
authors.initials: M
authors.lastname: Cazzola
authors.forename: Francesca
authors.initials: F
authors.lastname: Ferrari
authors.forename: Maurizio
authors.initials: M
authors.lastname: Ferrari
authors.forename: Paolo
authors.initials: P
authors.lastname: Arosio
mesh_terms: D020121:5' Untranslated Regions; D002851:Chromatography, High Pressure Liquid; D004587:Electrophoresis, Agar Gel; D005293:Ferritins / Q000097:blood* / Q000235:genetics*; D006801:Humans; D035925:Iron-Regulatory Proteins / Q000235:genetics; D009154:Mutation; D012313:RNA; D012045:Regulatory Sequences, Nucleic Acid
chemical_list: D020121:5' Untranslated Regions; D035925:Iron-Regulatory Proteins; D012313:RNA; D005293:Ferritins