title: Single molecule sequencing of free DNA from maternal plasma for noninvasive trisomy 21 detection.
abstract: BACKGROUND
  Noninvasive fetal aneuploidy detection by use of free DNA from maternal plasma has recently been shown to be achievable by whole genome shotgun sequencing. The high-throughput next-generation sequencing platforms previously tested use a PCR step during sample preparation, which results in amplification bias in GC-rich areas of the human genome. To eliminate this bias, and thereby experimental noise, we have used single molecule sequencing as an alternative method.
  
  
  METHODS
  For noninvasive trisomy 21 detection, we performed single molecule sequencing on the Helicos platform using free DNA isolated from maternal plasma from 9 weeks of gestation onwards. Relative sequence tag density ratios were calculated and results were directly compared to the previously described Illumina GAII platform.
  
  
  RESULTS
  Sequence data generated without an amplification step show no GC bias. Therefore, with the use of single molecule sequencing all trisomy 21 fetuses could be distinguished more clearly from euploid fetuses.
  
  
  CONCLUSIONS
  This study shows for the first time that single molecule sequencing is an attractive and easy to use alternative for reliable noninvasive fetal aneuploidy detection in diagnostics. With this approach, previously described experimental noise associated with PCR amplification, such as GC bias, can be overcome.
authors.affiliation: Center for Human and Clinical Genetics, Laboratory for Diagnostic Genome Analysis, Leiden University Medical Center, Leiden, the Netherlands.
authors.forename: Jessica M E
authors.initials: JM
authors.lastname: van den Oever
authors.forename: Sahila
authors.initials: S
authors.lastname: Balkassmi
authors.forename: E Joanne
authors.initials: EJ
authors.lastname: Verweij
authors.forename: Maarten
authors.initials: M
authors.lastname: van Iterson
authors.forename: Phebe N
authors.initials: PN
authors.lastname: Adama van Scheltema
authors.forename: Dick
authors.initials: D
authors.lastname: Oepkes
authors.forename: Jan M M
authors.initials: JM
authors.lastname: van Lith
authors.forename: Mariëtte J V
authors.initials: MJ
authors.lastname: Hoffer
authors.forename: Johan T
authors.initials: JT
authors.lastname: den Dunnen
authors.forename: Egbert
authors.initials: E
authors.lastname: Bakker
authors.forename: Elles M J
authors.initials: EM
authors.lastname: Boon
mesh_terms: D004247:DNA / Q000097:blood / Q000235:genetics*; D004314:Down Syndrome / Q000175:diagnosis*; D005260:Female; D005333:Fetus; D006801:Humans; D008297:Male; D016133:Polymerase Chain Reaction; D011247:Pregnancy; D011261:Pregnancy Trimester, First; D012189:Retrospective Studies; D017422:Sequence Analysis, DNA / Q000379:methods
chemical_list: D004247:DNA