title: Assessing the fidelity of ancient DNA sequences amplified from nuclear genes.
abstract: To date, the field of ancient DNA has relied almost exclusively on mitochondrial DNA (mtDNA) sequences. However, a number of recent studies have reported the successful recovery of ancient nuclear DNA (nuDNA) sequences, thereby allowing the characterization of genetic loci directly involved in phenotypic traits of extinct taxa. It is well documented that postmortem damage in ancient mtDNA can lead to the generation of artifactual sequences. However, as yet no one has thoroughly investigated the damage spectrum in ancient nuDNA. By comparing clone sequences from 23 fossil specimens, recovered from environments ranging from permafrost to desert, we demonstrate the presence of miscoding lesion damage in both the mtDNA and nuDNA, resulting in insertion of erroneous bases during amplification. Interestingly, no significant differences in the frequency of miscoding lesion damage are recorded between mtDNA and nuDNA despite great differences in cellular copy numbers. For both mtDNA and nuDNA, we find significant positive correlations between total sequence heterogeneity and the rates of type 1 transitions (adenine --> guanine and thymine --> cytosine) and type 2 transitions (cytosine --> thymine and guanine --> adenine), respectively. Type 2 transitions are by far the most dominant and increase relative to those of type 1 with damage load. The results suggest that the deamination of cytosine (and 5-methyl cytosine) to uracil (and thymine) is the main cause of miscoding lesions in both ancient mtDNA and nuDNA sequences. We argue that the problems presented by postmortem damage, as well as problems with contamination from exogenous sources of conserved nuclear genes, allelic variation, and the reliance on single nucleotide polymorphisms, call for great caution in studies relying on ancient nuDNA sequences.
authors.affiliation: Ancient DNA and Evolution Group, Centre for Ancient Genetics, Niels Bohr Institute, University of Copenhagen, Denmark.
authors.forename: Jonas
authors.initials: J
authors.lastname: Binladen
authors.forename: Carsten
authors.initials: C
authors.lastname: Wiuf
authors.forename: M Thomas P
authors.initials: MT
authors.lastname: Gilbert
authors.forename: Michael
authors.initials: M
authors.lastname: Bunce
authors.forename: Ross
authors.initials: R
authors.lastname: Barnett
authors.forename: Greger
authors.initials: G
authors.lastname: Larson
authors.forename: Alex D
authors.initials: AD
authors.lastname: Greenwood
authors.forename: James
authors.initials: J
authors.lastname: Haile
authors.forename: Simon Y W
authors.initials: SY
authors.lastname: Ho
authors.forename: Anders J
authors.initials: AJ
authors.lastname: Hansen
authors.forename: Eske
authors.initials: E
authors.lastname: Willerslev
mesh_terms: D000818:Animals; D002467:Cell Nucleus / Q000235:genetics*; D004272:DNA, Mitochondrial / Q000737:chemistry* / Q000235:genetics*; D005580:Fossils; D008322:Mammals / Q000235:genetics; D008969:Molecular Sequence Data; D046108:Palaeognathae / Q000235:genetics; D016133:Polymerase Chain Reaction / Q000379:methods*; D017422:Sequence Analysis, DNA
chemical_list: D004272:DNA, Mitochondrial