title: Nonradioactive multiplex PCR screening strategy for the simultaneous detection of multiple low-density lipoprotein receptor gene mutations.
abstract: We have developed a rapid, nonradioactive screening test enabling the simultaneous analysis of three low-density lipoprotein receptor (LDLR) gene mutations (D154N, D206E, and V408M), which together account for familial hypercholesterolemia (FH) in approximately 90% of the South African Afrikaner population. The assay is designed so that FH patients, negative for these founder-related mutations (found in descendants of European settlers), subsequently can be screened for unknown mutations in the mutation-rich exon 4 of the LDLR gene. Our screening assay consists of two steps: (1) multiplex allele-specific PCR amplification of exons 4 and 9, and (2) simultaneous analysis of single- and double-strand conformational polymorphisms in exon 4 by vertical electrophoresis on low cross-linked polyacrylamide gels. The simplicity, specificity, and versatility of the multiplex assay makes it an ideal system for routine screening of FH mutations in large population samples.
authors.affiliation: Department of Human Genetics, Faculty of Medicine, University of Stellenbosch, Tygerberg, South Africa.
authors.forename: M J
authors.initials: MJ
authors.lastname: Kotze
authors.forename: L
authors.initials: L
authors.lastname: Theart
authors.forename: M
authors.initials: M
authors.lastname: Callis
authors.forename: A V
authors.initials: AV
authors.lastname: Peeters
authors.forename: R
authors.initials: R
authors.lastname: Thiart
authors.forename: E
authors.initials: E
authors.lastname: Langenhoven
mesh_terms: D001483:Base Sequence; D004247:DNA / Q000235:genetics / Q000302:isolation & purification; D017931:DNA Primers; D004591:Electrophoresis, Polyacrylamide Gel; D005060:Europe / Q000208:ethnology; D005091:Exons; D006801:Humans; D006937:Hypercholesterolemia / Q000235:genetics*; D008969:Molecular Sequence Data; D017354:Point Mutation; D016133:Polymerase Chain Reaction / Q000379:methods*; D011110:Polymorphism, Genetic; D018807:Polymorphism, Single-Stranded Conformational; D011973:Receptors, LDL / Q000235:genetics*; D017384:Sequence Deletion; D013019:South Africa
chemical_list: D017931:DNA Primers; D011973:Receptors, LDL; D004247:DNA