title: Epigenome-wide DNA methylation assay reveals placental epigenetic markers for noninvasive fetal single-nucleotide polymorphism genotyping in maternal plasma.
abstract: BACKGROUND
  The use of DNA methylation difference between maternal blood cell and fetal (placental) DNA is one of the main areas of interest for the development of fetal epigenetics markers in maternal plasma.
  
  
  METHODS
  We employed a methylation array (HumanMethylation450 array, Illumina, Inc.) to identify novel biomarkers that are specially hypermethylated in placental DNA versus maternal blood cells in a genome-wide basis. Validation by bisulfite genomic sequencing was performed and the priority was given to potential targets that harbor differential methylated CpG sites overlapped with at least two methylation-sensitive restriction enzyme (MSRE) recognizing sites, as well as one polymorphic single-nucleotide polymorphism (SNP), within a short DNA stretch. Three candidate regions of PSMB8, SKI, and CHST11 gene were selected for developing a preliminary polymerase chain reaction assay with MSRE digestion of maternal plasma DNA. SNP genotypes were confirmed by direct sequencing.
  
  
  RESULTS
  We identified 2944 and 5218 fetal-specific hypermethylated CpG sites in the first- and third-trimester placenta, respectively, of which 2613 were overlapped, suggesting a consistency of differential methylation during the whole pregnancy. The array results were confirmed by bisulfite genomic sequencing. The preliminary tests in maternal plasma showed that postdigestion hypermathylated versions of these candidate molecules were detectable only in pregnant women. We further revealed that methylated targets in maternal plasma possessed the fetal SNP genotypes.
  
  
  CONCLUSIONS
  The present studies systematically identified hypermethylated sites in fetal tissues and preliminarily demonstrated that some of the fetal epigenetic markers that contain informative SNPs have great potential for noninvasive fetal genetic diagnosis.
authors.affiliation: Department of Forensic Medicine, Zhongshan School of Medicine, Sun Yat-sen University, Guangzhou, China.
authors.forename: Xueling
authors.initials: X
authors.lastname: Ou
authors.forename: Huan
authors.initials: H
authors.lastname: Wang
authors.forename: Dongyang
authors.initials: D
authors.lastname: Qu
authors.forename: Yongzhen
authors.initials: Y
authors.lastname: Chen
authors.forename: Jun
authors.initials: J
authors.lastname: Gao
authors.forename: Hongyu
authors.initials: H
authors.lastname: Sun
mesh_terms: D018899:CpG Islands; D019175:DNA Methylation; D044127:Epigenesis, Genetic; D005260:Female; D005333:Fetus / Q000378:metabolism; D005819:Genetic Markers; D005820:Genetic Testing; D060005:Genotyping Techniques / Q000379:methods; D006801:Humans; D020411:Oligonucleotide Array Sequence Analysis; D010920:Placenta / Q000378:metabolism*; D020641:Polymorphism, Single Nucleotide; D011247:Pregnancy; D011296:Prenatal Diagnosis / Q000379:methods*
chemical_list: D005819:Genetic Markers