title: Detection of increased amounts of cell-free fetal DNA with short PCR amplicons.
abstract: OBJECTIVE
  A digital PCR approach has recently been suggested to detect greater amounts of cell-free fetal DNA in maternal plasma than conventional real-time quantitative PCR (qPCR). Because the digital qPCR approach uses shorter PCR amplicons than the real-time qPCR assay, we investigated whether a real-time qPCR assay appropriately modified for such short amplicons would improve the detection of cell-free fetal DNA.
  
  
  METHODS
  We developed a novel universal-template (UT) real-time qPCR assay that was specific for the DYS14 sequence on Y chromosome and had a short amplicon size of 50 bp. We examined this "short" assay with 50 maternal plasma samples and compared the results with those for a conventional real-time qPCR assay of the same locus but with a longer amplicon (84 bp).
  
  
  RESULTS
  Qualitatively, both assays detected male cell-free fetal DNA with the same specificity and detection capability. Quantitatively, however, the new UT real-time qPCR assay for shorter amplicons detected, on average, almost 1.6-fold more cell-free fetal DNA than the conventional real-time qPCR assay with longer amplicons.
  
  
  CONCLUSIONS
  The use of short PCR amplicons improves the detection of cell-free fetal DNA. This feature may prove useful in attempts to detect cell-free fetal DNA under conditions in which the amount of template is low, such as in samples obtained early in pregnancy.
authors.affiliation: Department of Biomedicine, University Women's Hospital, Basel, Switzerland.
authors.forename: Aleksandra
authors.initials: A
authors.lastname: Sikora
authors.forename: Bernhard G
authors.initials: BG
authors.lastname: Zimmermann
authors.forename: Corinne
authors.initials: C
authors.lastname: Rusterholz
authors.forename: Daniella
authors.initials: D
authors.lastname: Birri
authors.forename: Varaprasad
authors.initials: V
authors.lastname: Kolla
authors.forename: Olav
authors.initials: O
authors.lastname: Lapaire
authors.forename: Irene
authors.initials: I
authors.lastname: Hoesli
authors.forename: Vivian
authors.initials: V
authors.lastname: Kiefer
authors.forename: Laird
authors.initials: L
authors.lastname: Jackson
authors.forename: Sinuhe
authors.initials: S
authors.lastname: Hahn
mesh_terms: D004247:DNA / Q000097:blood*; D005260:Female; D005333:Fetus; D006801:Humans; D008297:Male; D008431:Maternal-Fetal Exchange; D016133:Polymerase Chain Reaction / Q000379:methods*; D011247:Pregnancy; D012189:Retrospective Studies
chemical_list: D004247:DNA