title: Single-Stranded DNA Library Preparation Preferentially Enriches Short Maternal DNA in Maternal Plasma.
abstract: Recent studies have suggested that single-stranded DNA (ssDNA) library preparation can enrich short DNA species from the plasma of healthy individuals, cancer patients, and transplant recipients. Based on previous observations that fetal DNA molecules in the maternal plasma are shorter than maternal DNA molecules, ssDNA library preparation may potentially enrich fetal DNA and provide substantial improvement in noninvasive prenatal testing.
  
  
  
  We tested this hypothesis by comparing the maternal plasma DNA sequencing results using 2 types of ssDNA library preparation methods and a standard double-stranded DNA (dsDNA) library method using samples from first- and third-trimester pregnancies. We also evaluated the performance of ssDNA and dsDNA library methods in the noninvasive prenatal detection of trisomy 21 from maternal plasma.
  
  
  
  Short DNA species were significantly enriched in ssDNA libraries. However, contrary to previous speculation, no significant enrichment was observed in the overall fetal fraction in maternal plasma collected in the first trimester. Our use of an ssDNA library did not reduce the variation in chromosomal representation when compared with a standard dsDNA library in the first-trimester plasma samples. ssDNA libraries also showed inferior performance in the noninvasive prenatal detection of trisomy 21 from maternal plasma. Detailed fetal fraction analysis using size-fractionated Y chromosome sequences and fetal-specific single-nucleotide polymorphisms (SNPs) revealed an unexpected finding that short maternal DNA was preferentially enriched over short fetal DNA in an ssDNA library irrespective of GC content.
  
  
  
  Our findings have shown that ssDNA library preparation preferentially enriches short maternally derived DNA in maternal plasma.
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 SAR, China.|Departments of Chemical Pathology and.
authors.forename: Joaquim S L
authors.initials: JSL
authors.lastname: Vong
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 SAR, China.|Departments of Chemical Pathology and.
authors.forename: Jason C H
authors.initials: JCH
authors.lastname: Tsang
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 SAR, China.|Departments of Chemical Pathology and.
authors.forename: Peiyong
authors.initials: P
authors.lastname: Jiang
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 SAR, China.|Departments of Chemical Pathology and.
authors.forename: Wing-Shan
authors.initials: WS
authors.lastname: Lee
authors.affiliation: Obstetrics and Gynaecology, The Chinese University of Hong Kong, Shatin, New Territories, Hong Kong SAR, China.
authors.forename: Tak Yeung
authors.initials: TY
authors.lastname: Leung
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 SAR, China.|Departments of Chemical Pathology and.
authors.forename: K C Allen
authors.initials: KCA
authors.lastname: Chan
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 SAR, China.|Departments of Chemical Pathology and.
authors.forename: Rossa W K
authors.initials: RWK
authors.lastname: Chiu
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 SAR, China; loym@cuhk.edu.hk.|Departments of Chemical Pathology and.
authors.forename: Y M Dennis
authors.initials: YMD
authors.lastname: Lo
mesh_terms: D004247:DNA / Q000097:blood* / Q000737:chemistry / Q000235:genetics; D004277:DNA, Single-Stranded; D005260:Female; D015723:Gene Library; D006801:Humans; D008297:Male; D008967:Molecular Biology / Q000592:standards*; D008969:Molecular Sequence Data; D011247:Pregnancy
chemical_list: D004277:DNA, Single-Stranded; D004247:DNA