Development of methods for the characterization and retrieval of damaged DNA from ancient biological material
Over course of the last decade, the field of ancient DNA has been transformed by the advent of highthroughput sequencing. In specimens with exceptional DNA preservation, this allowed whole nuclear genomes of extinct organisms to be sequenced and analyzed. To deal with material with suboptimal DNA preservation and high levels of external modern DNA contamination as is common in intensively handled museum specimens, significant challenges remain. In this thesis I developed laboratory and in silico methods to make highly contaminated biological material amenable to genetic research and explore the structure and decay mechanisms of ancient DNA.
Chapter 1 illustrates the development of a library preparation method for high-throughput sequencing that selects for DNA molecules containing uracils, a damaged base typically found in ancent DNA, in order to enrich authentic ancient molecules against a background of modern contamination. The method was applied to Neanderthal samples from Gibraltar demonstrating its suitability for recovering DNA sequences from material that shows very poor DNA preservation and that is particularly strongly contaminated with modern human DNA.
Chapter 2 describes a new method to reconstruct the native double-stranded DNA molecules and overhang structures in DNA isolated from a Neanderthal specimen by combining deep sequencing of low-input single-stranded DNA libraries with in silico sequence matching. I analyzed patterns of nucleotide substitutions and base composition separately in the different structures found and showed also that the method can be applied to modern sources of fragmented DNA, specifically to cell-free DNA isolated from present-day blood samples.
| Identifer | oai:union.ndltd.org:DRESDEN/oai:qucosa:de:qucosa:74736 |
| Date | 03 May 2021 |
| Creators | Bokelmann, Lukas |
| Contributors | Universität Leipzig |
| Source Sets | Hochschulschriftenserver (HSSS) der SLUB Dresden |
| Language | English, German |
| Detected Language | English |
| Type | info:eu-repo/semantics/publishedVersion, doc-type:doctoralThesis, info:eu-repo/semantics/doctoralThesis, doc-type:Text |
| Rights | info:eu-repo/semantics/openAccess |
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