Nucleic Acids Research Advance Access originally published online on August 22, 2007
Nucleic Acids Research 2007 35(17):5717-5728; doi:10.1093/nar/gkm588
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Nucleic Acids Research, 2007, Vol. 35, No. 17 5717-5728
© 2007 The Author(s)
This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/2.0/uk/) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.
Molecular Biology |
Novel high-resolution characterization of ancient DNA reveals C > U-type base modification events as the sole cause of post mortem miscoding lesions
1Australian Centre for Ancient DNA, School of Earth and Environmental Sciences, University of Adelaide, Adelaide, SA 5005, Australia, 2Henry Wellcome Ancient Biomolecules Centre, Department of Zoology, University of Oxford, South Parks Road, Oxford OX1 3PS, UK, 3National Institute of Toxicology and Forensic Science, Canary Islands delegation, Tenerife, Spain, 4School of Animal and Microbial Sciences, University of Reading, Reading RG6 6AG, UK and 5Department of Archaeology, University of Durham, South Road, Durham DH1 3L, UK
*To whom correspondence should be addressed. Tel: +44 1457 858605; Email: paul.brotherton{at}virgin.net
Received February 25, 2007. Revised July 16, 2007. Accepted July 17, 2007.
Ancient DNA (aDNA) research has long depended on the power of PCR to amplify trace amounts of surviving genetic material from preserved specimens. While PCR permits specific loci to be targeted and amplified, in many ways it can be intrinsically unsuited to damaged and degraded aDNA templates. PCR amplification of aDNA can produce highly-skewed distributions with significant contributions from miscoding lesion damage and non-authentic sequence artefacts. As traditional PCR-based approaches have been unable to fully resolve the molecular nature of aDNA damage over many years, we have developed a novel single primer extension (SPEX)-based approach to generate more accurate sequence information. SPEX targets selected template strands at defined loci and can generate a quantifiable redundancy of coverage; providing new insights into the molecular nature of aDNA damage and fragmentation. SPEX sequence data reveals inherent limitations in both traditional and metagenomic PCR-based approaches to aDNA, which can make current damage analyses and correct genotyping of ancient specimens problematic. In contrast to previous aDNA studies, SPEX provides strong quantitative evidence that C > U-type base modifications are the sole cause of authentic endogenous damage-derived miscoding lesions. This new approach could allow ancient specimens to be genotyped with unprecedented accuracy.
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