title: Targeted massively parallel sequencing of maternal plasma DNA permits efficient and unbiased detection of fetal alleles.
abstract: BACKGROUND
  Massively parallel sequencing has recently been used in noninvasive prenatal diagnosis. The current costs of this technology are still relatively expensive, however, and sample throughput is still relatively low when it is used as a molecular diagnostic tool. Rather than nonselectively sequencing the genome, target enrichment provides a logical approach for more efficient and cost-effective massively parallel sequencing because it increases the proportion of informative data from the targeted region(s). Existing applications of targeted sequencing have mainly been qualitative analyses of genomic DNA. In this study, we investigated its applicability in enriching selected genomic regions from plasma DNA and the quantitative performance of this approach.
  
  
  METHODS
  DNA was extracted from plasma samples collected from 12 pregnant women carrying female fetuses. The SureSelect Target Enrichment System (Agilent Technologies) was used to enrich for exons on chromosome X. Plasma DNA libraries with and without target enrichment were analyzed by massively parallel sequencing. Genomic DNA samples of the mother and fetus for each case were genotyped by microarray.
  
  
  RESULTS
  For the regions targeted by the enrichment kit, the mean sequence coverage of the enriched samples was 213-fold higher than that of the nonenriched samples. Maternal and fetal DNA molecules were enriched evenly. After target enrichment, the coverage of fetus-specific alleles within the targeted region increased from 3.5% to 95.9%.
  
  
  CONCLUSIONS
  Targeted sequencing of maternal plasma DNA permits efficient and unbiased detection of fetal alleles at genomic regions of interest and is a powerful method for measuring the proportion of fetal DNA in a maternal plasma sample.
authors.affiliation: Centre for Research into Circulating Fetal Nucleic Acids, Li Ka Shing Institute of Health Sciences, The Chinese University of Hong Kong, Shatin, New Territories, Hong Kong.
authors.forename: Gary J W
authors.initials: GJ
authors.lastname: Liao
authors.forename: Fiona M F
authors.initials: FM
authors.lastname: Lun
authors.forename: Yama W L
authors.initials: YW
authors.lastname: Zheng
authors.forename: K C Allen
authors.initials: KC
authors.lastname: Chan
authors.forename: Tak Y
authors.initials: TY
authors.lastname: Leung
authors.forename: Tze K
authors.initials: TK
authors.lastname: Lau
authors.forename: Rossa W K
authors.initials: RW
authors.lastname: Chiu
authors.forename: Y M Dennis
authors.initials: YM
authors.lastname: Lo
mesh_terms: D000483:Alleles; D004247:DNA / Q000097:blood / Q000235:genetics*; D005260:Female; D005333:Fetus; D006801:Humans; D020411:Oligonucleotide Array Sequence Analysis; D020641:Polymorphism, Single Nucleotide; D011247:Pregnancy; D011264:Pregnancy Trimesters; D011296:Prenatal Diagnosis / Q000379:methods*; D017422:Sequence Analysis, DNA
chemical_list: D004247:DNA