Novel high-resolution characterization of ancient DNA reveals C > U-type base modification events as the sole cause of post mortem miscoding lesions.
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.
Brotherton
Paul
P
Australian Centre for Ancient DNA, School of Earth and Environmental Sciences, University of Adelaide, Adelaide, SA 5005, Australia. paul.brotherton@virgin.net
Endicott
Phillip
P
Sanchez
Juan J
JJ
Beaumont
Mark
M
Barnett
Ross
R
Austin
Jeremy
J
Cooper
Alan
A
D000241:Adenosine / Q000737:chemistry; D000818:Animals; D003596:Cytosine / Q000737:chemistry; D004247:DNA / Q000737:chemistry; D004249:DNA Damage; D053938:DNA Fragmentation; D017931:DNA Primers; D005580:Fossils; D006147:Guanine / Q000737:chemistry; D006801:Humans; D021141:Nucleic Acid Amplification Techniques / Q000379:methods*; D016133:Polymerase Chain Reaction; D017422:Sequence Analysis, DNA; D019914:Taq Polymerase; D013698:Templates, Genetic; D014498:Uracil / Q000737:chemistry
D017931:DNA Primers; D014498:Uracil; D006147:Guanine; D003596:Cytosine; D004247:DNA; D019914:Taq Polymerase; D000241:Adenosine
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