title: Discovery of Rare Haplotypes by Typing Millions of Single-Molecules with Bead Emulsion Haplotyping (BEH).
abstract: Characterizing polymorphisms on single molecules renders the phase of different alleles, and thus, haplotype information. Here, we describe a high-throughput method to genotype hundreds-of thousands single molecules in parallel using bead-emulsion haplotyping (BEH). Haplotyping via BEH is an emulsion-PCR-based method that was adapted to amplify multiple DNA fragments on paramagnetic, microscopic beads within a compartment formed by an aqueous-oil emulsion. This generates beads covered by thousands of clonal copies from several polymorphic regions of an initial DNA molecule that are then genotyped with fluorescently labeled probes. With BEH, up to three different polymorphisms (or more if several polymorphisms are within an amplicon) can be typed within a fragment of several kilobases in a singleexperiment, rendering haplotype information of a very large number of initial single molecules. The high throughput and digital nature of the method makes it ideal to quantify rare haplotypes or to assess the haplotype diversity in complex samples.
authors.affiliation: Institute of Biophysics, Johannes Kepler University, Gruberstraße 40, 4020, Linz, Austria.
authors.forename: Elisabeth
authors.initials: E
authors.lastname: Palzenberger
authors.affiliation: Institute of Biophysics, Johannes Kepler University, Gruberstraße 40, 4020, Linz, Austria.
authors.forename: Ronja
authors.initials: R
authors.lastname: Reinhardt
authors.affiliation: Cambridge Advanced Imaging Centre, University of Cambridge, Cambridge, UK.
authors.forename: Leila
authors.initials: L
authors.lastname: Muresan
authors.affiliation: Institute of Biophysics, Johannes Kepler University, Gruberstraße 40, 4020, Linz, Austria.
authors.forename: Barbara
authors.initials: B
authors.lastname: Palaoro
authors.affiliation: Institute of Biophysics, Johannes Kepler University, Gruberstraße 40, 4020, Linz, Austria. Irene.tiemann@jku.at.
authors.forename: Irene
authors.initials: I
authors.lastname: Tiemann-Boege
mesh_terms: D000483:Alleles; D005838:Genotype; D006239:Haplotypes / Q000235:genetics*; D006801:Humans; D016133:Polymerase Chain Reaction; D020641:Polymorphism, Single Nucleotide / Q000235:genetics
keywords: Allelic phase; Bead-emulsion haplotyping (BEH); Digital PCR; Emulsion PCR; High-throughput haplotyping; Single-molecule amplification