title: Tumour necrosis factor receptor superfamily member 6 gene mutation detection by denaturing high-performance liquid chromatography.
abstract: Denaturing high-performance liquid chromatography (DHPLC) was evaluated as a tool for diagnostic screening of polymorphisms in the tumour necrosis factor receptor superfamily member 6 (TNFRSF6) also known as CD95, Apo-1 or Fas gene. Exons 1-9 of the TNFRSF6 gene were amplified from genomic DNA of 38 individuals, of which three were known to carry mutations in the TNFRSF6 gene. The TNFRSF6 gene amplicons were analysed for heterozygosity by DHPLC. Samples that displayed heterozygous variation by DHPLC were further analysed by sequencing. Comparison of DHPLC analysis with sequencing results showed an overall 100% concordance for samples in which heterozygosity was detected by DHPLC. Importantly, DHPLC was in all cases able to demonstrate the presence or absence of mutations in exon 9 encoding the death domain of the TNFRSF6 gene, which have been implied as the most frequent genetic cause of autoimmune lymphoproliferative syndrome. Comparison of DHPLC analysis with sequencing results showed an overall 100% concordance for samples in which heterozygosity was detected by DHPLC. In conclusion, DHPLC is a suitable method for the detection of genetic variation in the TNFRSF6 gene.
authors.affiliation: Institute of Immunology, Rikshospitalet University Hospital, Oslo, Norway. g.e.etokebe@labmed.uio.no
authors.forename: G E
authors.initials: GE
authors.lastname: Etokebe
authors.forename: T G
authors.initials: TG
authors.lastname: Abrahamsen
authors.forename: B
authors.initials: B
authors.lastname: Bogen
authors.forename: A
authors.initials: A
authors.lastname: Spurkland
mesh_terms: D001483:Base Sequence; D002675:Child, Preschool; D002851:Chromatography, High Pressure Liquid; D004252:DNA Mutational Analysis; D017931:DNA Primers; D005820:Genetic Testing; D006801:Humans; D007154:Immune System Diseases / Q000175:diagnosis* / Q000235:genetics*; D008969:Molecular Sequence Data; D009154:Mutation; D016133:Polymerase Chain Reaction; D011110:Polymorphism, Genetic; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D019014:fas Receptor / Q000235:genetics*
chemical_list: D017931:DNA Primers; D019014:fas Receptor