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Non-radioactive DNA diagnosis for the fragile X syndrome in mentally retarded Japanese males

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      Abstract

      A rapid screening test was developed to detect CGG repeat expansion of the FMR-1 gene causing the fragile X syndrome by a non-radioisotope PCR technique. A biotin-labeled primer was initially used and the biotin-labeled PCR product was detected by means of chemiluminescence. The normal PCR product of around 300 bp was not created in the abnormal FMR-1 gene sample with this method. Four positive samples were found among those from 226 mentally retarded males, but the CGG repeat expansion was shown on Southern blot analysis in only one sample. To eliminate false-positive samples, a hybridization method involving a biotin-labeled (CGG)5 oligonucleotide was developed for the PCR product and the CGG repeat expansion could be detected. Finally, 256 mentally retarded males in Japan were examined and only 2 abnormal samples were detected. The prevalence of this abnormality was less than 1%, which is relatively lower than those reported previously.

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      Reference

        • Turner G.
        • Robinson H.
        • Laing S.
        • Purris-Smith S.
        Preventive screening for the fragile X syndrome.
        N Eng J Med. 1986; 315: 607-609
        • Oberle I.
        • Rousseau F.
        • Heitz D.
        • et al.
        Instability of a 550-base pair DNA segment and abnormal methylation in fragile X syndrome.
        Science. 1991; 252: 1097-1102
        • Bell M.V.
        • Hirst M.C.
        • Nakahori Y.
        • et al.
        Physical mapping across the fragile X: hypermethylation and clinical expression of the fragile X syndrome.
        Cell. 1991; 64: 861-866
        • Yu S.
        • Pritchard M.
        • Kremer E.
        • et al.
        Fragile X genotype characterized by an unstable region of DNA.
        Science. 1991; 252: 1179-1181
        • Fu Y.H.
        • Kuhl D.P.
        • Pizzuti A.
        • et al.
        Variation of the CGG repeat at the fragile X site results in genetic instability: resolution of the Sherman paradox.
        Cell. 1991; 67: 1047-1058
        • Kremer E.J.
        • Pritchard M.
        • Lynch M.
        • et al.
        Mapping of DNA instability at the fragile X to a trinucleotide repeat sequence p(CGG)n.
        Science. 1991; 252: 1711-1714
        • Sherman S.L.
        • Jacobs P.A.
        • Morton N.E.
        • et al.
        Further segregation analysis of the fragile (X) syndrome with special reference to transmitting males.
        Hum Genet. 1985; 69: 289-299
        • Richards R.I.
        • Sutherland G.R.
        Dynamic mutations: a new class of mutations causing human disease.
        Cell. 1992; 70: 709-712
        • Caskey C.T.
        • Pizzuti A.
        • Fu Y.-H.
        • Fenwick J.R.G.
        • Nelson D.L.
        Triplet repeat mutations in human disease.
        Science. 1992; 256: 784-789
        • Rousseau F.
        • Heitz D.
        • Biancalana V.
        • et al.
        Direct diagnosis by DNA analysis of the fragile X syndrome of mental retardation.
        N Engl J Med. 1991; 325: 1673-1681
        • Nakahori Y.
        • Knight S.J.L.
        • Holland J.
        • et al.
        Molecular heterogeneity of the fragile X syndrome.
        Nucleic Acids Res. 1991; 19: 4355-4359
        • Snow K.
        • Doud L.K.
        • Hagerman R.
        • Pergolizzi R.G.
        • Erster S.H.
        • Thibodeau S.N.
        Analysis of a CGG sequence at the FMR-1 locus in fragile X families and in the general population.
        Am J Hum Genet. 1993; 53: 1217-1228
        • Erster S.H.
        • Brown W.T.
        • Goonewardena P.
        • Dobkin C.S.
        • Jenkins E.C.
        • Pergolizzi R.G.
        Polymerase chain reaction analysis of fragile X mutations.
        Hum Genet. 1992; 90: 55-61
        • Pergolizzi R.
        • Erster S.H.
        • Goonewardena P.
        • Brown W.T.
        Detection of full fragile X mutation.
        Lancet. 1992; 339: 271-272
        • Brown W.T.
        • Houck G.E.J.
        • Jeziorowska A.
        • et al.
        Rapid fragile X carrier screening and prenatal diagnosis using a nonradioactive PCR test.
        JAMA. 1993; 270: 1569-1575
        • Hashimoto K.
        • Takeuchi A.
        • Ieshima A.
        Fragile X syndrome: the report of 4 patients from a family (in Japanese).
        Tottori Igaku Zasshi (Yonago). 1992; 20: 340-342
        • Higuchi R.
        Simple and raid preparation of samples for PCR.
        in: Erlich H.A. PCR technology: Principles and applications for DNA amplification. Stockton Press, New York1989: 31-38
        • Beck S.
        • O'Keeffe T.
        • Coull J.M.
        • Koster H.
        Chemiluminescent detection of DNA: application for DNA sequencing and hybridization.
        Nucleic Acids Res. 1989; 17: 5115-5123
      1. 2nd edn. Molecular cloning: A laboratory manual. Cold Spring Harbor University Press, Cold Spring Harbor1989
        • Engler-Blum G.
        • Meier M.
        • Frank J.
        • Muller G.A.
        Reduction of background problems in non-radioactive northern and southern blot analyses enables higher sensitivity than32P-based hybridizations.
        Anal Biochem. 1993; 210: 235-244
        • De Boulle K.
        • Verkerk A.J.R.E.
        • Vits L.
        • et al.
        A point mutation in the FMR-1 gene associated with fragile X mental retardation.
        Nature Genet. 1993; 3: 31-35
        • Tarleton J.
        • Richie R.
        • Schwartz C.
        • Rao K.
        • Aylsworth A.S.
        • Lachiewicz A.
        An extensive de novo deletion removing FMR1 in a patient with mental retardation and the fragile X syndrome phenotype.
        Hum Mol Genet. 1993; 2: 1973-1974
        • Sutherland G.R.
        Heritable fragile sites on human chromosomes. VIII. Preliminary population cytogenetic data on the folic acid-sensitive fragile sites.
        Am J Hum Genet. 1982; 34: 452-458
        • Froster-Iskenius U.
        • Felsch G.
        • Schirren C.
        • Schwinger E.
        Screening for fra(X)(q) in a population of mentally retarded males.
        Hum Genet. 1983; 63: 153-157
        • Blomquist H.K.
        • Gustavson K.H.
        • Holmgren G.
        • Nordenson I.
        • Palson-Stae U.
        Fragile X syndrome in mildly mentally retarded children in a Northern Swedish county. A prevalence study.
        Clin Genet. 1983; 24: 393-398
        • Kirkilionis A.
        • Sergovich F.
        • Pozsonyi J.
        Use of testicular volume as a cytogenetic screening criterion.
        Am J Hum Genet. 1983; 35: 138A
        • Jacobs P.A.
        • Mayer M.
        • Matsuura J.
        • Rhoads F.
        • Yee S.C.
        A cytogenetic study of a population of mentally retarded males with special reference to the marker (X) syndrome.
        Hum Genet. 1983; 63: 139-148
        • Bundey S.
        • Webb T.P.
        • Thake A.
        • Todd J.
        A community study of severe mental retardation in the West Midlands and the importance of the fragile X chromosome in its aetiology.
        J Med Genet. 1985; 22: 258-266
        • Arinami T.
        • Kondo I.
        • Nakajima S.
        Frequency of the fragile X syndrome in Japanese mentally retarded males.
        Hum Genet. 1986; 73: 309-312
        • Turner G.
        • Robinson H.
        • Laing S.
        • et al.
        Population screening for fragile X.
        Lancet. 1992; 336: 1210-1213
        • Jacobs P.A.
        • Bullman H.
        • Macpherson J.
        • et al.
        Population studies of the fragile X: a molecular approach.
        J Med Genet. 1993; 30: 454-459
        • Hofstee Y.
        • Arinami T.
        • Hamaguchi H.
        Comparison between the cytogenetic test for fragile X and the molecular analysis of the FMR-1 gene in Japanese mentally retarded individuals.
        Am J Med Genet. 1994; 51: 466-470
        • Wang Q.
        • Green E.
        • Barnicoat A.
        • et al.
        Cytogenetic versus DNA diagnosis in routine referrals for fragile X syndrome.
        Lancet. 1993; 342: 1025-1026
        • Knight S.J.L.
        • Flannery A.V.
        • Hirst M.C.
        • et al.
        Trinucleotide repeat amplification and hypermethylation of a CpG island in FRAXE mental retardation.
        Cell. 1993; 74: 127-134
        • Rousseau F.
        • Heitz D.
        • Tarleton J.
        • et al.
        A multicenter study on genotype-phenotype correlations in the fragile X syndrome, using direct diagnosis with probe StB12.3: the first 2,253 cases.
        Am J Hum Genet. 1994; 55: 225-237
        • Arinami T.
        • Asano M.
        • Kobayashi K.
        • Yanagi H.
        • Hamaguchi H.
        Data on the CGG repeat at the fragile X site in the non-retarded Japanese population and family suggest the presence of a subgroup of normal alleles predisposing to mutate.
        Hum Genet. 1993; 92: 431-436
        • Richards R.I.
        • Holman K.
        • Friend K.
        • et al.
        Evidence of founder chromosomes in fragile X syndrome.
        Nature Genet. 1992; 1: 257-260
        • Kunst C.B.
        • Warren S.T.
        Cryptic and polar variation of the fragile X repeat could result in predisposing normal alleles.
        Cell. 1994; 77: 853-861
        • Hashimoto O.
        • Shimizu Y.
        • Kawasaki Y.
        Brief report: low frequency of the fragile X syndrome among Japanese autistic subjects.
        J Autism Dev Disord. 1993; 23: 201-209