title: [Methylation analysis of CpG island DNA of FMR1 gene in the fragile X syndrome].
abstract: OBJECTIVE
  To establish a method of methylation-sensitive restriction enzymes based quantitative PCR (MSRE-qPCR) for analysis of CpG island DNA of FMR1 gene, and to assess its value for molecular diagnosis of fragile X syndrome.
  
  
  METHODS
  Thirty boys with mental retardation and abnormal repeats of 5'(CGG)n in the FMR1 gene and 20 mothers were analyzed by conventional PCR screening. Eag I was used to digest genomic DNA, and qPCR was performed to amplify CpG island in the FMR1 gene using both undigested and digested templates. Raw Ct values were obtained through quantitative PCR amplification. The degree of CpG island methylation was calculated by 2 - U+0394 U+0394 Ct. The result of MSRE-qPCR was verified by Southern blotting. 30 healthy females and 30 healthy males were used as controls to optimize the established MSRE-qPCR method.
  
  
  RESULTS
  The ranges of 2 - U+0394 U+0394 Ct value for normal methylation, partial methylation and full methylation were determined. Among the 30 patients, 3 were found to have partial methylation of CpG island of the FMR1 gene, and 27 were found to have full methylation (3/30 results were verified by Southern blotting). Only 7 mothers were found abnormal methylation of CpG island of FMR1 gene, whilst the remaining 13 mothers were normal.
  
  
  CONCLUSIONS
  MSRE-qPCR is a quick and reliable method for quantitative analysis of CpG island methylation status in FMR1 gene, which may provide a new strategy for the diagnosis of fragile X syndrome.
authors: Department of Genetics and Metabolism, Zhejiang University, Hangzhou, Zhejiang, People's Republic of China.
authors: Ding-wen
authors: DW
authors: Wu
authors: Zhi-wei
authors: ZW
authors: Zhu
authors: Zheng-yan
authors: ZY
authors: Zhao
authors: Yi-ping
authors: YP
authors: Qu
authors: Jian-bin
authors: JB
authors: Yang
mesh_terms: D018899:CpG Islands; D019175:DNA Methylation; D005260:Female; D051860:Fragile X Messenger Ribonucleoprotein 1 / Q000235:genetics*; D005600:Fragile X Syndrome / Q000175:diagnosis / Q000235:genetics*; D006801:Humans; D008297:Male; D012737:Sex Factors
chemical_list: D051860:Fragile X Messenger Ribonucleoprotein 1; C497683:FMR1 protein, human