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  • Review Article
  • Published:

Reconstructing ancient genomes and epigenomes

Nature Reviews Genetics volume 16, pages 395–408 (2015)Cite this article

Subjects

Key Points

  • High-throughput sequencing technologies have revolutionized ancient DNA (aDNA) research by enabling the reconstruction of whole-genome sequences from traces of short and extremely degraded DNA fragments.

  • DNA preservation is highly variable across samples and environments, as well as within single archaeological remains. The current temporal range for whole-genome sequencing covers the past million years.

  • DNA extracts from ancient material are generally metagenomic assemblages that include the DNA from the host and its associated microorganisms, as well as from a range of environmental microorganisms that colonize the sample after its death.

  • Various molecular approaches have been developed to improve access to aDNA in samples and reduce the sequencing costs of paleogenomics. These include extraction methods tailored to ultrashort DNA fragments, target enrichment for library inserts annealing to a panel of nucleic acid probes, and library building procedures targeting each DNA strand individually or incorporating only the most damaged DNA fragments.

  • Analyses of aDNA are prone to contamination by modern DNA molecules, which generally show limited degradation and fragmentation. Therefore, ruling out contamination — for example, exploiting patterns of DNA degradation and monitoring the heterozygosity levels observed at haploid loci — represents the cornerstone of every aDNA study.

  • DNA degradation reactions taking place post-mortem introduce mutation and depth-of-coverage patterns in the sequence data that can be exploited to authenticate paleogenomes and reconstruct genome-wide nucleosome and methylation maps.

Abstract

Research involving ancient DNA (aDNA) has experienced a true technological revolution in recent years through advances in the recovery of aDNA and, particularly, through applications of high-throughput sequencing. Formerly restricted to the analysis of only limited amounts of genetic information, aDNA studies have now progressed to whole-genome sequencing for an increasing number of ancient individuals and extinct species, as well as to epigenomic characterization. Such advances have enabled the sequencing of specimens of up to 1 million years old, which, owing to their extensive DNA damage and contamination, were previously not amenable to genetic analyses. In this Review, we discuss these varied technical challenges and solutions for sequencing ancient genomes and epigenomes.

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Figure 1: Major advances in ancient genomics.
Figure 2: Typical ancient DNA molecules.
Figure 3: Constructing ancient DNA libraries.
Figure 4: Enriching DNA libraries for ancient inserts.
Figure 5: Tracking ancient nucleosome and methylation maps.

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References

  1. Higuchi, R., Bowman, B., Freiberger, M., Ryder, O. A. & Wilson, A. C. DNA sequences from the quagga, an extinct member of the horse family. Nature 312, 282–284 (1984).

    Article  CAS  PubMed  Google Scholar 

  2. Hagelberg, E., Sykes, B. & Hedges, R. Ancient bone DNA amplified. Nature 342, 485 (1989).

    Article  CAS  PubMed  Google Scholar 

  3. Pääbo, S. Ancient DNA: extraction, characterization, molecular cloning, and enzymatic amplification. Proc. Natl Acad. Sci. USA 86, 1939–1943 (1989).

    Article  PubMed  Google Scholar 

  4. Rasmussen, M. et al. Ancient human genome sequence of an extinct Paleo-Eskimo. Nature 463, 757–762 (2010). This study takes advantage of the relative absence of environmental microorganisms within ancient hairs to characterize the first high-quality ancient human genome.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  5. Miller, W. et al. Sequencing the nuclear genome of the extinct woolly mammoth. Nature 456, 387–390 (2008).

    Article  CAS  PubMed  Google Scholar 

  6. Green, R. E. et al. A draft sequence of the Neandertal genome. Science 328, 710–722 (2010). This paper reports the first draft genome of an archaic hominin and many methodological developments that are still commonly used for characterizing and analysing ancient genomes.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  7. Bos, K. I. et al. A draft genome of Yersinia pestis from victims of the Black Death. Nature 478, 506–510 (2011). This paper reports the first genome isolated from an ancient pathogenic bacterium, confirming the Black Death as a plague epidemic. It revealed that no derived variant is unique to the medieval strain, suggesting that non-genetic factors enhanced the virulence of the pathogen.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  8. Martin, M. D. et al. Reconstructing genome evolution in historic samples of the Irish potato famine pathogen. Nat. Commun. 4, 2172 (2013).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  9. Schuenemann, V. J. et al. Genome-wide comparison of medieval and modern Mycobacterium leprae. Science 341, 179–183 (2013).

    Article  CAS  PubMed  Google Scholar 

  10. Bos, K. I. et al. Pre-Columbian mycobacterial genomes reveal seals as a source of New World human tuberculosis. Nature 514, 494–497 (2014).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  11. Devault, A. M. et al. Second-pandemic strain of Vibrio cholera from the Philadelphia cholera outbreak. N. Engl. J. Med. 370, 334–340 (2014).

    Article  CAS  PubMed  Google Scholar 

  12. Devault, A. M. et al. Ancient pathogen DNA in archaeological samples detected with a microbial detection array. Sci. Rep. 4, 4245 (2014).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  13. Wagner, D. M. et al. Yersinia pestis and the Plague of Justinian 541–543 AD: a genomic analysis. Lancet Infect. Dis. 14, 319–326 (2014).

    Article  PubMed  Google Scholar 

  14. Rasmussen, M. et al. An Aboriginal Australian genome reveals separate human dispersals into Asia. Science 334, 94–98 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  15. Keller, A. et al. New insights into the Tyrolean Iceman's origin and phenotype as inferred by whole-genome sequencing. Nat. Commun. 3, 698 (2012).

    Article  CAS  PubMed  Google Scholar 

  16. Meyer, M. et al. A high-coverage genome sequence from an archaic Denisovan individual. Science 338, 222–226 (2012). This paper describes a novel method for constructing aDNA libraries using ssDNA templates, which enabled the characterization of the Denisovan genome at a quality rivalling that of modern genomes, starting from only minute amounts of DNA extracts.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  17. Orlando, L. et al. Recalibrating Equus evolution using the genome sequence of an early Middle Pleistocene horse. Nature 499, 74–78 (2013). This study takes advantage of both second-generation (high-throughput, and library- and amplification-dependent) and third-generation (high-throughput, and library- and amplification-independent) sequencing technologies to present the oldest genome sequence hitherto characterized: that of an ~700,000-year-old horse.

    Article  CAS  PubMed  Google Scholar 

  18. Gamba, C. et al. Genome flux and stasis in a five millennium transect of European prehistory. Nat. Commun. 5, 5257 (2014).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  19. Jónsson, H. et al. Speciation with gene flow in equids despite extensive chromosomal plasticity. Proc. Natl Acad. Sci. USA 111, 18655–18660 (2014).

    Article  CAS  PubMed  Google Scholar 

  20. Malaspinas, A. S. et al. Two ancient human genomes reveal Polynesian ancestry among the indigenous Botocudos of Brazil. Curr. Biol. 24, R1035–R1037 (2014).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  21. Olalde, I. et al. Derived immune and ancestral pigmentation alleles in a 7,000-year-old Mesolithic European. Nature 507, 225–228 (2014).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  22. Prüfer, K. et al. The complete genome sequence of a Neanderthal from the Altai Mountains. Nature 505, 43–49 (2014).

    Article  CAS  PubMed  Google Scholar 

  23. Raghavan, M. et al. The genetic prehistory of the New World Arctic. Science 345, 1255832 (2014).

    Article  CAS  PubMed  Google Scholar 

  24. Raghavan, M. et al. Upper Paleolithic Siberian genome reveals dual ancestry of Native Americans. Nature 505, 87–91 (2014).

    Article  CAS  PubMed  Google Scholar 

  25. Rasmussen, M. et al. The genome of a Late Pleistocene human from a Clovis burial site in western Montana. Nature 506, 225–229 (2014).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  26. Schubert, M. et al. Prehistoric genomes reveal the genetic foundation and cost of horse domestication. Proc. Natl Acad. Sci. USA 111, E5661–E5669 (2014).

    Article  CAS  PubMed  Google Scholar 

  27. Seguin-Orlando, A. et al. Genomic structure in Europeans dating back at least 36,200 years. Science 346, 1113–1118 (2014).

    Article  CAS  PubMed  Google Scholar 

  28. Ramirez, O. et al. Genome data from a sixteenth century pig illuminate modern breed relationships. Heredity 114, 175–184 (2015).

    Article  CAS  PubMed  Google Scholar 

  29. Schroeder, H. et al. Genome-wide ancestry of 17th century enslaved Africans from the Caribbean. Proc. Natl Acad. Sci. USA 112, 3669–3673 (2015).

    Article  CAS  PubMed  Google Scholar 

  30. Metzker, M. L. Sequencing technologies — the next generation. Nat. Rev. Genet. 11, 31–46 (2010).

    Article  CAS  PubMed  Google Scholar 

  31. Dabney, J. et al. Complete mitochondrial genome sequence of a Middle Pleistocene cave bear reconstructed from ultrashort DNA fragments. Proc. Natl Acad. Sci. USA 110, 15758–15763 (2013).

    Article  CAS  PubMed  Google Scholar 

  32. Meyer, M. et al. A mitochondrial genome sequence of a hominin from Sima de los Huesos. Nature 505, 403–406 (2014).

    Article  CAS  PubMed  Google Scholar 

  33. Gokhman, D. et al. Reconstructing the DNA methylation maps of the Neandertal and the Denisovan. Science 344, 523–527 (2014).

    Article  CAS  PubMed  Google Scholar 

  34. Pedersen, J. S. et al. Genome-wide nucleosome map and cytosine methylation levels of an ancient human genome. Genome Res. 24, 454–466 (2014). This study exploits DNA degradation patterns in HTS data sets to characterize, for the first time, genome-wide nucleosome and methylation maps from an ancient human and infer ancient gene expression levels and the age at death of the individual.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  35. Ermini, L., Der Sarkissian, C., Willerslev, E. & Orlando, L. Major transitions in human evolution revisited: a tribute to ancient DNA. J. Hum. Evol. 79, 4–20 (2015).

    Article  PubMed  Google Scholar 

  36. Shapiro, B. & Hofreiter, M. A paleogenomic perspective on evolution and gene function: new insights from ancient DNA. Science 343, 1236573 (2014).

    Article  CAS  PubMed  Google Scholar 

  37. Orlando, L. & Cooper, A. Using ancient DNA to understand evolutionary and ecological processes. Ann. Rev. Ecol. Evol. Syst. 45, 573–598 (2014).

    Article  Google Scholar 

  38. Orlando, L. & Willerslev, E. An epigenetic window into the past? Science 345, 511–512 (2014).

    Article  CAS  PubMed  Google Scholar 

  39. Höss, M., Jaruga, P., Zastawny, T. H., Dizdaroglu, M. & Pääbo, S. DNA damage and DNA sequence retrieval from ancient tissues. Nucleic Acids Res. 24, 1304–1307 (1996).

    Article  PubMed  PubMed Central  Google Scholar 

  40. Hansen, A. J., Willerslev, E., Wiuf, C., Mourier, T. & Arctander, P. Statistical evidence for miscoding lesions in ancient DNA templates. Mol. Biol. Evol. 18, 262–265 (2001).

    Article  CAS  PubMed  Google Scholar 

  41. Hofreiter, M., Jaenicke, V., Serre, D., von Haeseler, A. & Pääbo, S. DNA sequences from multiple amplifications reveal artifacts induced by cytosine deamination in ancient DNA. Nucleic Acids Res. 29, 4793–4799 (2001).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  42. Stiller, M. et al. Patterns of nucleotide misincorporations during enzymatic amplification and direct large-scale sequencing of ancient DNA. Proc. Natl Acad. Sci. USA 103, 13578–13584 (2006).

    Article  CAS  PubMed  Google Scholar 

  43. Gilbert, M. T. et al. Recharacterization of ancient DNA miscoding lesions: insights in the era of sequencing-by-synthesis. Nucleic Acids Res. 35, 1–10 (2007).

    Article  CAS  PubMed  Google Scholar 

  44. Briggs, A. et al. Patterns of damage in genomic DNA sequences from a Neandertal. Proc. Natl Acad. Sci. USA 104, 14616–14621 (2007). This study characterizes typical nucleotide misincorporation and fragmentation patterns using HTS data from aDNA extracts, which have been subsequently used as essential authentication criteria.

    Article  CAS  PubMed  Google Scholar 

  45. Orlando, L. et al. True single-molecule DNA sequencing of a Pleistocene horse bone. Genome Res. 21, 1705–1719 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  46. Sawyer, S. et al. Temporal patterns of nucleotide misincorporations and DNA fragmentation in ancient DNA. PLoS ONE 7, e34131 (2012).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  47. Overballe-Petersen, S., Orlando, L. & Willerslev, E. Next-generation sequencing offers new insights into DNA degradation. Trends Biotechnol. 30, 364–368 (2012).

    Article  CAS  PubMed  Google Scholar 

  48. Jónsson, H. et al. mapDamage2.0: fast approximate Bayesian estimates of ancient DNA damage parameters. Bioinformatics 29, 1682–1684 (2013).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  49. Hansen, A. J. et al. Crosslinks rather than strand breaks determine access to ancient DNA sequences from frozen sediments. Genet. 173, 1175–1179 (2006).

    Article  CAS  Google Scholar 

  50. Heyn, P. et al. Road blocks on paleogenomes — polymerase extension profiling reveals the frequency of blocking lesions in ancient DNA. Nucleic Acids Res. 38, e161 (2010).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  51. Poinar, H. N., Kuch, M., McDonald, G., Martin, P. & Pääbo, S. Nuclear gene sequences from a late Pleistocene sloth coprolithe. Curr. Biol. 13, 1150–1152 (2003).

    Article  CAS  PubMed  Google Scholar 

  52. Poinar, H. N. et al. Metagenomics to paleogenomics: large-scale sequencing of mammoth DNA. Science 311, 393–394 (2006). This study reports the first genetic analysis of ancient specimens based on a HTS technology, paving the way for whole-genome sequencing from ancient specimens.

    Article  CAS  Google Scholar 

  53. Allentoft, M. E. et al. The half-life of DNA in bone: measuring decay kinetics in 158 dated fossils. Proc. Biol. Sci. 279, 4724–4733 (2012).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  54. Smith, C. I., Chamberlain, A. T., Riley, M. S., Stringer, C. & Collins, M. J. The thermal history of human fossils and the likelihood of successful DNA amplification. J. Hum. Evol. 45, 203–217 (2003).

    Article  PubMed  Google Scholar 

  55. Schwarz, C. et al. New insights from old bones: DNA preservation and degradation in permafrost preserved mammoth remains. Nucleic Acids Res. 37, 3215–2129 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  56. Ginolhac, A. et al. Improving the performance of true single molecule sequencing for ancient DNA. BMC Genomics 13, 177 (2012).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  57. Der Sarkissian, C. et al. Shotgun microbial profiling of fossil remains. Mol. Ecol. 23, 1780–1798 (2014).

    Article  CAS  PubMed  Google Scholar 

  58. Damgaard, P. et al. Improving access to endogenous DNA in ancient bones and teeth. BioRxiv http://dx.doi.org/10.1101/014985 (2015).

  59. Salamon, M., Tuross, N., Arensburg, B. & Weiner, S. Relatively well preserved DNA is present in the crystal aggregates of fossil bones. Proc. Natl Acad. Sci. USA 102, 13783–13788 (2005).

    Article  CAS  PubMed  Google Scholar 

  60. Adler, C. J., Haak, W., Donlon, D. & Cooper, A. Survival and recovery of DNA from ancient teeth and bones. J. Archaeol. Sci. 38, 956–964 (2011).

    Article  Google Scholar 

  61. Seguin-Orlando, A. et al. Ligation bias in illumina next-generation DNA libraries: implications for sequencing ancient genomes. PLoS ONE 8, e78575 (2013).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  62. Dabney, J. & Meyer, M. Length and GC-biases during sequencing library amplification: a comparison of various polymerase-buffer systems with ancient and modern DNA sequencing libraries. Biotechniques 87–94 (2012).

  63. Young, A. L. et al. A new strategy for genome assembly using short sequence reads and reduced representation libraries. Genome Res. 20, 249–256 (2010).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  64. Seguin-Orlando, A. et al. Amplification of TruSeq ancient DNA libraries with AccuPrime Pfx: consequences on nucleotide misincorporation and methylation patterns. STAR 1, STAR2015112054892315Y.0000000005 (2015).

    Article  Google Scholar 

  65. Star, B. et al. Palindromic sequence artifacts generated during next generation sequencing library preparation from historic and ancient DNA. PLoS ONE 9, e89676 (2014).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  66. Gansauge, M. T. & Meyer, M. Single-stranded DNA library preparation for the sequencing of ancient or damaged DNA. Nat. Protoc. 8, 737–748 (2013).

    Article  CAS  PubMed  Google Scholar 

  67. Gilbert, M. T. et al. Whole-genome shotgun sequencing of mitochondria from ancient hair shafts. Science 317, 1927–1930 (2007).

    Article  CAS  PubMed  Google Scholar 

  68. Gansauge, M. T. & Meyer, M. Selective enrichment of damaged DNA molecules for ancient genome sequencing. Genome Res. 24, 1543–1549 (2014).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  69. Briggs, A. et al. Targeted retrieval and analysis of five Neandertal mtDNA genomes. Science 325, 318–321 (2009).

    Article  CAS  PubMed  Google Scholar 

  70. Maricic, T., Whitten, M. & Pääbo, S. Multiplexed DNA sequence capture of mitochondrial genomes using PCR products. PLoS ONE 5, e14004 (2010).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  71. Haak, W. et al. Massive migration from the steppe was a source for Indo-European languages in Europe. Nature http://dx.doi.org/10.1038/nature14317 (2015).

  72. Rohland, N., Harney, E., Mallick, S., Nordenfelt, S. & Reich, D. Partial uracil–DNA–glycosylase treatment for screening of ancient DNA. Phil. Trans. R. Soc. B 370, 20130624 (2015).

    Article  CAS  PubMed  Google Scholar 

  73. Burbano, H. A. et al. Targeted investigation of the Neandertal genome by array-based sequence capture. Science 328, 723–725 (2010). This paper reports the first characterization of an ancient exome using target enrichment approaches on microarrays.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  74. Fu, Q. et al. A revised timescale for human evolution based on ancient mitochondrial genomes. Curr. Biol. 23, 553–559 (2013).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  75. Vilstrup, J. T. et al. Mitochondrial phylogenomics of modern and ancient equids. PLoS ONE 8, e55950 (2013).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  76. Castellano, S. et al. Patterns of coding variation in the complete exomes of three Neandertals. Proc. Natl Acad. Sci. USA 111, 6666–6671 (2014).

    Article  CAS  PubMed  Google Scholar 

  77. Fu, Q. et al. DNA analysis of an early modern human form Tianyuan Cave, China. Proc. Natl Acad. Sci. USA 110, 2223–2227 (2013). This paper describes a target enrichment procedure exploiting millions of DNA probes cleaved from user-designed DNA microarrays to characterize the almost complete sequence of the non-repetitive fraction of chromosome 21 for an ~40,000-year-old human.

    Article  CAS  PubMed  Google Scholar 

  78. Carpenter, M. L. et al. Pulling out the 1%: whole-genome capture for the targeted enrichment of ancient DNA sequencing libraries. Am. J. Hum. Genet. 93, 852–864 (2013). This paper reports the first whole-genome target enrichment method, which makes use of self-generated RNA probes. The method substantially reduces the operational cost of target enrichment and allows genetic analyses of specimens with only minute amounts of aDNA templates.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  79. Enk, J. M. et al. Ancient whole genome enrichment using baits built from modern DNA. Mol. Biol. Evol. 31, 1292–1295 (2014).

    Article  CAS  PubMed  Google Scholar 

  80. Avila-Arcos, C. et al. Comparative performance of two whole-genome capture methodologies on ancient DNA Illumina libraries. Methods Ecol. Evol. http://dx.doi.org/10.1111/2041-210X.12353 (2015).

  81. Briggs, A. et al. Removal of deaminated cytosines and detection of in vivo methylation in ancient DNA. Nucleic Acids Res. 38, e87 (2010). This paper presents an enzymatic procedure based on the treatment of DNA extracts with USER mix, which can considerably reduce the sequencing error rate of ancient genomes by limiting the effect of nucleotide misincorporations at damaged sites.

    Article  CAS  PubMed  Google Scholar 

  82. Mason, V. C., Li, G., Helgen, K. M. & Murphy, W. J. Efficient cross-species capture hybridization and next-generation sequencing of mitochondrial genomes from noninvasively sampled museum specimens. Genome Res. 21, 1695–1704 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  83. Zhang, H. et al. Morphological and genetic evidence for early Holocene cattle management in northeastern China. Nat. Commun. 4, 2755 (2013).

    Article  CAS  PubMed  Google Scholar 

  84. Fabre, P. H. et al. Rodents of the Caribbean: origin and diversification of hutias unraveled by next-generation museomics. Biol. Lett. http://dx.doi.org/10.1098/rsbl.2014.0266 (2014).

  85. Foote, A. D. et al. Tracking niche variation over millennial timescales in sympatric killer whale lineages. Proc. Biol. Sci. 280, 20131481 (2013).

    Article  PubMed  PubMed Central  Google Scholar 

  86. Schuenemann, V. J. et al. Targeted enrichment of ancient pathogens yielding the pPCP1 plasmid of Yersinia pestis from victims of the Black Death. Proc. Natl Acad. Sci. USA 108, E746–E452 (2011).

    Article  CAS  PubMed  Google Scholar 

  87. Kircher, M., Sawyer, S. & Meyer, M. Double indexing overcomes inaccuracies in multiplex sequencing on the Illumina platform. Nucleic Acids Res. 40, e3 (2012).

    Article  CAS  PubMed  Google Scholar 

  88. Avila-Arcos, M. C. et al. Application and comparison of large-scale solution-based DNA capture-enrichment methods on ancient DNA. Sci. Rep. 1, 73 (2011).

    Article  CAS  Google Scholar 

  89. Hodges, E. et al. Hybrid selection of discrete genomic intervals on custom-designed microarrays for massively parallel sequencing. Nat. Protoc. 4, 960–974 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  90. Bos, K. I. et al. Parallel detection of ancient pathogens via array-based DNA capture. Phil. Trans. R. Soc. B 370, 20130375 (2015).

    Article  CAS  PubMed  Google Scholar 

  91. Schubert, M. et al. Characterization of ancient and modern genomes by SNP detection and phylogenomic and metagenomic analysis using PALEOMIX. Nat. Protoc. 9, 1056–1082 (2013). This paper presents a fully automated pipeline performing all sequence analyses associated with re-sequencing genomic projects, phylogenomic inference and metagenomic profiling. It is applicable to both modern and ancient sequence data sets.

    Article  CAS  Google Scholar 

  92. Lindgreen, S. AdapterRemoval: easy cleaning of next-generation sequencing reads. BMC Res. Notes 5, 337 (2012).

    Article  PubMed  PubMed Central  Google Scholar 

  93. Li, H. & Durbin, R. Fast and accurate short read alignment with Burrows–Wheeler transform. Bioinformatics 25, 1754–1760 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  94. Langmead, B. & Salzberg, S. L. Fast gapped-read alignment with Bowtie 2. Nat. Methods 9, 357–359 (2012).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  95. McKenna, A. et al. The genome analysis toolkit: a MapReduce framework for analyzing next-generation DNA sequencing data. Genome Res. 20, 1297–1303 (2010).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  96. Kozlov, A. M., Aberer, A. J. & Stamatakis, A. ExaML version 3: a tool for phylogenomic analyses on supercomputers. Bioinformatics http://dx.doi.org/10.1093/bioinformatics/btv184 (2015).

  97. Segata, N. et al. Metagenomic microbial community profiling using unique clade-specific marker genes. Nat. Methods 9, 811–814 (2012).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  98. Schubert, M. et al. Improving ancient DNA read mapping against modern reference genomes. BMC Genomics 13, 178 (2012).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  99. Kerpedjev, P., Frellsen, J., Lindgreen, S. & Krogh, A. Adaptable probabilistic mapping of short reads using position specific scoring matrices. BMC Bioinformatics 15, 100 (2014).

    Article  Google Scholar 

  100. Ginolhac, A., Rasmussen, M., Gilbert, M. T., Willerslev, E. & Orlando, L. mapDamage: testing for damage patterns in ancient DNA sequences. Bioinformatics 27, 2153–2155 (2011).

    Article  CAS  PubMed  Google Scholar 

  101. Skoglund, P. et al. Separating endogenous ancient DNA from modern day contamination in a Siberian Neandertal. Proc. Natl Acad. Sci. USA 111, 2229–2234 (2014).

    Article  CAS  PubMed  Google Scholar 

  102. Lindgreen, S., Krogh, A. & Pedersen, J. S. SNPest: a probabilistic graphical model for estimating genotypes. BMC Res. Notes 7, 698 (2014).

    Article  PubMed  PubMed Central  Google Scholar 

  103. Skoglund, P. et al. Origins and genetic legacy of Neolithic farmers and hunter-gatherers in Europe. Science 336, 466–469 (2012).

    Article  CAS  PubMed  Google Scholar 

  104. García-Garcerà, M. et al. Fragmentation of contaminant and endogenous DNA in ancient samples determined by shotgun sequencing; prospects for human paleogenomics. PLoS ONE 6, e24161 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  105. Sánchez-Quinto, F. et al. Genomic affinities of two 7,000-year-old Iberian hunter-gatherers. Curr. Biol. 22, 1494–1499 (2012).

    Article  CAS  PubMed  Google Scholar 

  106. Green, R. E. et al. The Neandertal genome and ancient DNA authenticity. EMBO J. 28, 2494–2502 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  107. Reich, D. et al. Genetic history of an archaic hominin group from Denisova Cave in Siberia. Nature 468, 1053–1060 (2010).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  108. Llamas, B. et al. High-resolution analysis of cytosine methylation in ancient DNA. PLoS ONE 7, e30226 (2012).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  109. Smith, O. et al. Genomic methylation patterns in archaeological barley show de-methylation as a time-dependent diagenetic process. Sci. Rep. 4, 5559 (2014).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  110. d'Abbadie, M. et al. Molecular breeding of polymerases for amplification of ancient DNA. Nat. Biotech. 25, 939–943 (2007).

    Article  CAS  Google Scholar 

  111. da Fonseca, R. R. et al. The origin and evolution of maize in the American Southwest. Nat. Plants 1, 14003 (2015).

    Article  PubMed  Google Scholar 

  112. Pickrell, J. K. & Reich, D. Toward a new history and geography of human genes informed by ancient DNA. Trends Genet. 30, 377–389 (2014).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  113. Fordyce, S. L. et al. Deep sequencing of RNA from ancient maize kernels. PLoS ONE 8, e50961 (2013).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  114. Allaby, R. G. et al. Using archaeogenomic and computational approaches to unravel the history of local adaptation in crops. Phil. Trans. R. Soc. B 370, 20130377 (2015).

    Article  CAS  PubMed  Google Scholar 

  115. Cappellini, E. et al. Proteomic analysis of a Pleistocene mammoth femur reveals more than one hundred ancient bone proteins. 11, 917–926 (2012).

  116. Cappellini, E., Collins, M. J. & Gilbert, M. T. Unlocking ancient protein palimpsests. Science 343, 1320–1322 (2014).

    Article  CAS  PubMed  Google Scholar 

  117. Warinner, C. et al. Direct evidence of milk consumption from ancient human dental calculus. Sci. Rep. 4, 7104 (2014).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  118. Adler, C. J. et al. Sequencing ancient calcified dental plaque shows changes in oral microbiota with dietary shifts of the Neolithic and Industrial revolutions. Nat. Genet. 45, 450–455 (2013).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  119. Warinner, C. et al. Pathogens and host immunity in the ancient human oral cavity. Nat. Genet. 46, 336–344 (2014).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  120. Shapiro, B. How to Clone a Mammoth — the Science of De-extinction (Princeton University Press, 2014).

    Google Scholar 

  121. Lazaridis, I. et al. Ancient human genomes suggest three ancestral populations for present-day Europeans. Nature 513, 409–413 (2014).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  122. Fu, Q. et al. Genome sequence of a 45,000-year-old modern human from western Siberia. Nature 514, 445–449 (2014).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  123. Krause, J. et al. The complete mitochondrial DNA genome of an unknown hominin from southern Siberia. Nature 464, 894–897 (2010).

    Article  CAS  PubMed  Google Scholar 

  124. Orlando, L. A. 400,000-year-old mitochondrial genome questions phylogenetic relationships amongst archaic hominins. Bioessays 36, 598–605 (2014).

    Article  CAS  PubMed  Google Scholar 

  125. Scally, A. & Durbin, R. Revising the human mutation rate: implications for understanding human evolution. Nat. Rev. Genet. 13, 745–753 (2012).

    Article  CAS  PubMed  Google Scholar 

  126. Alexander, D. H., Novembre, J. & Lange, K. Fast model-based estimation of ancestry in unrelated individuals. Genome Res. 19, 1655–1664 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  127. Malaspinas, A. S. et al. bammds: a tools for assessing the ancestry of low-depth whole-genome data using multidimensional scaling (MDS). Bioinformatics 30, 2962–2964 (2014).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  128. Skoglund, P., Sjodin, P., Skoglund, T., Lascoux, M. & Jakobsson, M. Investigating population history using temporal genetic differentiation. Mol. Biol. Evol. 31, 2516–2527 (2014).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  129. Durand, E. Y., Patterson, N., Reich, D. & Slatkin, M. Testing for ancient admixture between closely related populations. Mol. Biol. Evol. 28, 2239–2252 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  130. Patterson, N. et al. Ancient admixture in human history. Genetics. 192, 1065–1093 (2012).

    Article  PubMed  PubMed Central  Google Scholar 

  131. Eriksson, A. & Manica, A. Effect of ancient population structure on the degree of polymorphism shared between modern human populations and ancient hominins. Proc. Natl Acad. Sci. USA 109, 13956–13960 (2012).

    Article  CAS  PubMed  Google Scholar 

  132. Sankararaman, S., Patterson, N., Heng, L., Pääbo, S. & Reich, D. The date of interbreeding between Neandertals and modern humans. PLoS Genet. 8, e1002947 (2012).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

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Acknowledgements

This work was supported by the Danish Council for Independent Research, Natural Sciences (FNU, 4002-00152B and 0602-02383B); the Danish National Research Foundation (DNFR94); the Lundbeck Foundation (R52-5062); a Marie Curie Career Integration Grant (FP7 CIG-293845); and the “Chaires d'Attractivité 2014” IDEX, University of Toulouse, France.

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Authors and Affiliations

  1. Centre for GeoGenetics, Natural History Museum of Denmark, University of Copenhagen, Øster Voldgade 5–7, Copenhagen, 1350C, Denmark

    Ludovic Orlando, M. Thomas P. Gilbert & Eske Willerslev

  2. Université de Toulouse, University Paul Sabatier (UPS), Laboratoire AMIS, CNRS UMR 5288, 37 allées Jules Guesde, Toulouse, 31000, France

    Ludovic Orlando

  3. Department of Environment and Agriculture, Trace and Environmental DNA Laboratory, Curtin University, Perth, 6102, Western Australia, Australia

    M. Thomas P. Gilbert

Authors
  1. Ludovic Orlando
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Correspondence to Ludovic Orlando.

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Glossary

Osseous materials

Calcified animal tissues, such as bones and teeth.

Second-generation sequencing

High-throughput short-read DNA sequencing platforms that require library construction and thus modification of the DNA before sequencing. Most commonly represented by the Illumina, GS-FLX (454), ABI SOLiD and Ion Torrent series.

Resonance structures

Dynamic, alternative forms of molecular groups, such as nucleotide bases, that result from electron delocalization within the molecule.

Pre-digestion

Exposure of ancient calcified materials to a short initial digestion aimed at removing substantial fractions of exogenous contaminants.

454

The initial generation of GS-FLX sequencing platforms based on pyrosequencing, before their acquisition and renaming by Roche.

T/A ligation

A common DNA ligation technology that relies on complementary pairing of thymine and adenine overhangs at the 3′ ends of the adaptors and inserts to be ligated, respectively.

Shotgun sequencing

The sequencing of fragmented DNA in the absence of any selection strategy.

Primer extension capture

(PEC). An enrichment technology based on the ligation of short 5′-biotinylated oligonucleotides (including a 12-nucleotide-long spacer followed by a primer of 18–25 nucleotides that is designed to match a particular region of interest) to single-stranded target molecules. This is followed by a single round of polymerase-based extension so as to increase the length over which the molecules are hybridized.

Tiled probes

Probes that overlap in their positioning on the target so as to ensure that every target position is covered by more than one different probe.

Chimeric DNA libraries

Recombination between libraries containing different template molecules during library PCR amplification, resulting in hybrid (chimeric) sequences that do not represent true biological sequences.

Double-indexed DNA libraries

DNA libraries in which short (for example, 8 bp long) unique nucleotide indexes are incorporated within both adaptors used during library construction. Indexes are bordered by known sequences that serve to prime index sequencing reactions and also enable library attachment to the surface of the sequencing flow cell.

Mate pairs

Pairs of sequences derived from both ends of a DNA library.

Edit distance

The number of sequence mismatch counts between reads and targets.

Probabilistic aligners

Mapping algorithms that can accommodate non-uniform distributions of sequencing errors along reads, generally leading to improved alignments between reads and reference genomes.

Thermal age

The predicted time that it would have taken an archaeological sample to produce the observed degree of DNA degradation were the sample exposed to a constant temperature of 10 °C since deposition. Thermal age has been proposed to adjust the chronological age of a sample to its thermal history and to help in predicting the likelihood of DNA surviving in archaeological remains.

Haplotypes

The DNA sequences of haploid chromosomes.

Derived alleles

Alleles that are evolutionarily derived in a lineage of interest and that are not represented in an ancestral population or species.

Ancestral alleles

Alleles in the ancestral state before a mutation took place in a descending population, species or lineage.

Nearly fixed

Fixed alleles are those that are derived and present in all individuals in a descendent population or species. Nearly fixed alleles therefore represent those that are present in nearly all individuals (thus close to fixation, for example, showing allelic frequencies of 99% in the population).

Epialleles

Allelic variants showing identical genetic sequences but different epigenetic marks, such as different methylation patterns.

Ghost population

An unsampled population that exchanges migrants with other sampled populations and that can be identified based on admixture signatures left in descending populations.

Introgressive block lengths

Population admixture introduces a mosaic of ancestry blocks along the genome, the lengths of which decrease with each subsequent generation owing to recombination. Introgressive block lengths can therefore be exploited to determine the date of admixture events.

CTCF regions

Genomic regions targeted by CCCTC-binding factor (CTCF) and involved in regulating the three-dimensional structure of chromatin and transcription by mediating long-range interactions between genomic sequences.

Admixture

Interbreeding of individuals from multiple population origins, resulting in the introduction of DNA from one population into the genomes of a second population.

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Orlando, L., Gilbert, M. & Willerslev, E. Reconstructing ancient genomes and epigenomes. Nat Rev Genet 16, 395–408 (2015). https://doi.org/10.1038/nrg3935

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