文献_Screening for CGG Repeat Expansion in the FMR1 Gene by Melting Curve Analysis of Combined 5′ and 3′ Direct Triplet-Primed PCRs.pdfVIP

Clinical Chemistry 58:3 Molecular Diagnostics and Genetics 568–579 (2012) Screening for CGG Repeat Expansion in the FMR1 Gene by Melting Curve Analysis of Combined 5 and 3 Direct Triplet-Primed PCRs Clara R.L. Teo,1 Hai-Yang Law,2 Caroline G. Lee,3,4,5 and Samuel S. Chong1,6,7* BACKGROUND: CGG repeat expansions in the FMR1 assays minimizes the incidence of false-negative re- (fragile X mental retardation 1) gene are associated sults arising from repeat-flanking deletions. with fragile X syndrome, fragile X–associated trem- © 2011 American Association for Clinical Chemistry or/ataxia syndrome, and fragile X–associated pri- mary ovarian insufficiency. We evaluated the use of melting curve analysis (MCA) of triplet-primed PCR Fragile X syndrome (FXS)8 (OMIM 300624), the most (TP-PCR) assays as a rapid screening tool for the common heritable cause of mental retardation and the positive identification of expanded FMR1 alleles in second most frequent cause of mental retardation after men and women. Down syndrome, affects approximately 1 in 4000 males and 1 in 6000 females (1, 2). It is largely caused by METHODS: Both 5 - and 3-weighted direct TP-PCRs full-mutation (FM) CGG repeat expansions in the 5 (dTP-PCRs) were evaluated on 29 cell line– derived untranslated region of the FMR1 (fragile X mental re- DNA samples and 44 blinded clinical samples. The tardation 1) gene on the X chromosome, which lead to presence of expansions was identified by the melting hy