title: Amplification-free sequencing of cell-free DNA for prenatal non-invasive diagnosis of chromosomal aberrations.
abstract: Cell-free DNA has been used for fetal rhesus factor and sex determination, fetal aneuploidy screening, cancer diagnostics and monitoring, and other applications. However current methods of using cell free DNA require amplification, which leads to allelic dropout and bias especially when starting with small amounts of DNA. Here we describe an amplification-free method for sequencing of cell-free DNA, even from low levels of starting material. We evaluated this method in the context of prenatal diagnosis of fetal aneuploidy and compared it with a PCR-based library preparation method as well as a recently described method using unique molecular identifiers (UMI). All methods performed well, however coverage was increased by the amplification-free method and GC-induced bias was reduced by both the amplification-free method and the UMI method. Future diagnostic applications including whole genome sequencing of cell-free DNA will benefit from amplification-free sequencing.
authors.affiliation: Laboratory for Molecular Neurobiology, Department of Medical Biochemistry and Biophysics, Karolinska Institutet, Scheeles väg 1, SE-171 77 Stockholm, Sweden.
authors.forename: Kasper
authors.initials: K
authors.lastname: Karlsson
authors.affiliation: Clinical Genetics Unit, Department of Molecular Medicine and Surgery, Center for Molecular Medicine, Karolinska Institutet, Karolinska University Hospital Solna, 17176 Stockholm, Sweden.
authors.forename: Ellika
authors.initials: E
authors.lastname: Sahlin
authors.affiliation: Clinical Genetics Unit, Department of Molecular Medicine and Surgery, Center for Molecular Medicine, Karolinska Institutet, Karolinska University Hospital Solna, 17176 Stockholm, Sweden.
authors.forename: Erik
authors.initials: E
authors.lastname: Iwarsson
authors.affiliation: Karolinska Institutet, Department of Obstet Gynecol, Division of Clinical Science, Intervention and Technology, Stockholm, Sweden.
authors.forename: Magnus
authors.initials: M
authors.lastname: Westgren
authors.affiliation: Clinical Genetics Unit, Department of Molecular Medicine and Surgery, Center for Molecular Medicine, Karolinska Institutet, Karolinska University Hospital Solna, 17176 Stockholm, Sweden.
authors.forename: Magnus
authors.initials: M
authors.lastname: Nordenskjöld
authors.affiliation: Laboratory for Molecular Neurobiology, Department of Medical Biochemistry and Biophysics, Karolinska Institutet, Scheeles väg 1, SE-171 77 Stockholm, Sweden. Electronic address: Sten.linnarsson@ki.se.
authors.forename: Sten
authors.initials: S
authors.lastname: Linnarsson
mesh_terms: D000782:Aneuploidy; D005260:Female; D005820:Genetic Testing / Q000379:methods*; D006801:Humans; D016133:Polymerase Chain Reaction; D011247:Pregnancy; D011296:Prenatal Diagnosis / Q000379:methods*; D017422:Sequence Analysis, DNA / Q000379:methods*
keywords: Amplification-free; Bias; Cell-free DNA; Fetal aneuploidy; Karyotyping; Library preparation