title: Multiplex amplification of ancient DNA.
abstract: This method is designed to assemble long, continuous DNA sequences using minimal amounts of fragmented ancient DNA as template. This is achieved by a two-step approach. In the first step, multiple fragments are simultaneously amplified in a single multiplex reaction. Subsequently, each of the generated fragments is amplified individually using a single primer pair, in a standard simplex (monoplex) PCR. The ability to amplify multiple fragments simultaneously in the first step allows the generation of large amounts of sequence from rare template DNA, whereas the second nested step increases specificity and decreases amplification of contaminating DNA. In contrast to current protocols using many template-consuming simplex PCRs, the method described allows amplification of several kilobases of sequence in just one reaction. It thus combines optimal template usage with a high specificity and can be performed within a day.
authors.affiliation: Molecular Biochemistry, Institute of Biochemistry, Medical Faculty, University of Leipzig, Johannisallee 30, 04103 Leipzig, Germany.
authors.forename: Holger
authors.initials: H
authors.lastname: Römpler
authors.forename: Paul H
authors.initials: PH
authors.lastname: Dear
authors.forename: Johannes
authors.initials: J
authors.lastname: Krause
authors.forename: Matthias
authors.initials: M
authors.lastname: Meyer
authors.forename: Nadin
authors.initials: N
authors.lastname: Rohland
authors.forename: Torsten
authors.initials: T
authors.lastname: Schöneberg
authors.forename: Helen
authors.initials: H
authors.lastname: Spriggs
authors.forename: Mathias
authors.initials: M
authors.lastname: Stiller
authors.forename: Michael
authors.initials: M
authors.lastname: Hofreiter
mesh_terms: D001106:Archaeology / Q000379:methods; D004247:DNA / Q000032:analysis* / Q000235:genetics*; D007753:Laboratories; D010163:Paleontology / Q000379:methods; D016133:Polymerase Chain Reaction / Q000295:instrumentation / Q000379:methods*; D015203:Reproducibility of Results; D013698:Templates, Genetic; D013997:Time Factors
chemical_list: D004247:DNA