title: Factor IX gene mutations causing haemophilia B: comparison of SSC screening versus systematic DNA sequencing and diagnostic applications.
abstract: The search for mutations of the factor IX gene responsible for haemophilia B should nowadays be used routinely for the molecular diagnosis of this inherited disorder, i.e. carrier detection and prenatal diagnosis. A number of methodologies have been proposed, most of them being delicate or expensive. We have used a simple strategy based on a preliminary screening of eight factor IX gene fragments by single-strand conformation analysis (SSCA), followed by direct sequencing of fragments displaying an abnormal migration pattern. Carrier testing is then performed by use of an enzyme restriction site altered by the mutation or by the SSCA itself. By using this strategy we were able readily to identify the factor IX molecular defect of nine unrelated haemophilia B patients from southern France. We validated the efficiency and reliability of the SSC-based detection of mutations by sequencing all the polymerase chain reaction (PCR) fragments studied in the haemophilic patients. No other sequence alteration could be found except the one detected by SSC analysis. We conclude that this method can be advantageously used for diagnosis purposes in a routine laboratory involved in haemophilia B diagnosis and report nine previously undescribed haemophilia B families with their factor IX mutation.
authors.affiliation: Laboratoire de Biochimie Génétique, Centre Hospitalo-Universitaire et INSERM U249/CNRS UPR 9008, Montpellier, France.
authors.forename: P
authors.initials: P
authors.lastname: Aguilar-Martinez
authors.forename: M C
authors.initials: MC
authors.lastname: Romey
authors.forename: J F
authors.initials: JF
authors.lastname: Schved
authors.forename: J C
authors.initials: JC
authors.lastname: Gris
authors.forename: J
authors.initials: J
authors.lastname: Demaille
authors.forename: M
authors.initials: M
authors.lastname: Claustres
mesh_terms: D004252:DNA Mutational Analysis; D004277:DNA, Single-Stranded / Q000032:analysis* / Q000737:chemistry; D004587:Electrophoresis, Agar Gel; D005164:Factor IX / Q000032:analysis / Q000235:genetics*; D006580:Genetic Carrier Screening; D006239:Haplotypes; D002836:Hemophilia B / Q000175:diagnosis / Q000235:genetics*; D006801:Humans; D007124:Immunoenzyme Techniques; D009154:Mutation; D009690:Nucleic Acid Conformation; D016133:Polymerase Chain Reaction; D011110:Polymorphism, Genetic
chemical_list: D004277:DNA, Single-Stranded; D005164:Factor IX