title: A PCR-based test suitable for screening for fragile X syndrome among mentally retarded males.
abstract: Ever since the identification of the genetic cause of fragile X syndrome as the expansion of an unstable trinucleotide sequence, several diagnostic strategies have evolved from molecular studies. However, we still lack a simple test suitable for population screening. We have therefore developed a nonisotopic polymerase chain reaction (PCR)-based technique for the identification of fragile X full mutations among men, with easy visualization of the PCR products on silver-stained polyacrylamide gels. The technique consists of PCR amplification with primers that flank the trinucleotide repeats, with a product of 557 bp for the (CGG)29 allele. Conditions were established such that full mutations failed to amplify and were thus identified with 98% sensitivity compared with Southern blot analysis. To produce an indispensable internal control we added to the reaction a third primer, internal to this fragment, allowing the multiplex amplification of a monomorphic band corresponding to a CG-rich stretch 147 bp upstream of the polymorphic region. In trials involving 41 patients and 74 controls, the PCR-based test here described showed specificity of more than 98.6%, accuracy of 99% and a sensitivity of 98%. Thus, although not suitable for medical diagnosis, it constitutes a useful tool for screening for the fragile X syndrome in populations of mentally retarded males.
authors.affiliation: Departamento de Bioquímica, Universidade Federal de Minas Gerais, Belo Horizonte, Brazil.
authors.forename: L A
authors.initials: LA
authors.lastname: Haddad
authors.forename: R C
authors.initials: RC
authors.lastname: Mingroni-Netto
authors.forename: A M
authors.initials: AM
authors.lastname: Vianna-Morgante
authors.forename: S D
authors.initials: SD
authors.lastname: Pena
mesh_terms: D000483:Alleles; D001482:Base Composition; D001483:Base Sequence; D015139:Blotting, Southern / Q000379:methods; D018900:Dinucleotide Repeats / Q000235:genetics; D005260:Female; D051860:Fragile X Messenger Ribonucleoprotein 1; D005600:Fragile X Syndrome / Q000175:diagnosis* / Q000235:genetics*; D006580:Genetic Carrier Screening; D005820:Genetic Testing; D006801:Humans; D008607:Intellectual Disability / Q000235:genetics; D008297:Male; D008969:Molecular Sequence Data; D009030:Mosaicism; D009419:Nerve Tissue Proteins / Q000235:genetics; D016133:Polymerase Chain Reaction / Q000379:methods*; D016601:RNA-Binding Proteins; D012680:Sensitivity and Specificity; D018911:Trinucleotide Repeats / Q000235:genetics
chemical_list: D051860:Fragile X Messenger Ribonucleoprotein 1; D009419:Nerve Tissue Proteins; D016601:RNA-Binding Proteins; C497683:FMR1 protein, human