Field Of Ancient DNA Research Articles

A brief history of palaeogenomics: How a young discipline revolutionised the study of the past

In a few years, the field of ancient DNA has changed from an anecdotal and artisanal discipline to one of the most dynamic scientific fields, generating massive genomic data about hundreds of individuals from the past. These include from extinct hominins such as Neanderthals or Denisovans to prehistoric humans that inform us about the recent settlement of continents. Palaeogenomics provides direct information about time and space, adaptive and demographic aspects of human populations, and manifests complex patterns of past migrations that help us to understand the current diversity. The development of the discipline is a unique opportunity to establish collaborative links with archaeologists and anthropologists and create a truly multidisciplinary view of the study of the past

Evolutionary Patterns and Processes: Lessons from Ancient DNA.

Ever since its emergence in 1984, the field of ancient DNA has struggled to overcome the challenges related to the decay of DNA molecules in the fossil record. With the recent development of high-throughput DNA sequencing technologies and molecular techniques tailored to ultra-damaged templates, it has now come of age, merging together approaches in phylogenomics, population genomics, epigenomics, and metagenomics. Leveraging on complete temporal sample series, ancient DNA provides direct access to the most important dimension in evolution—time, allowing a wealth of fundamental evolutionary processes to be addressed at unprecedented resolution. This review taps into the most recent findings in ancient DNA research to present analyses of ancient genomic and metagenomic data.

Ancient DNA: the next generation - chapter and verse

As the field of ancient DNA (aDNA) enters its third decade, it is perhaps time to reflect on the amazing transformation that it has undergone. During the first two decades, analyses of aDNA were mainly focussed on mitochondrial and/or chloroplast DNA as a result of their multicopy abundance in the cell, making retrieval and reproducibility much easier. Study of mitochondrial DNA through time allows evolutionary relationships between species to be resolved, molecular clocks to be calibrated, the geographical origins of samples to be revealed, and the investigation of demographic histories. However, not until the advent of massive parallel sequencing [also know as second-generation sequencing and next-generation sequencing (NGS)] was possible to retrieve and study nuclear DNA on a more routine base. Ancient nuclear DNA can additionally be used to identify extinct phenotypes, assess the degree of admixture, and examine selection pressures. This is a short review of what has been, what may come, and how aDNA has influenced NGS. Although examples from archaeology are used to illustrate the impact of NGS technologies on the field, this approach has also been successfully applied to a range of disciplines, such as medicine and wildlife forensics.

Screening ancient tuberculosis with qPCR: challenges and opportunities.

The field of ancient DNA (aDNA) has rapidly accelerated in recent years as a result of new methods in next-generation sequencing, library preparation and targeted enrichment. Such research is restricted, however, by the highly variable DNA preservation within different tissues, especially when isolating ancient pathogens from human remains. Identifying positive candidate samples via quantitative PCR (qPCR) for downstream procedures can reduce reagent costs, increase capture efficiency and maximize the number of sequencing reads of the target. This study uses four qPCR assays designed to target regions within the Mycobacterium tuberculosis complex (MTBC) to examine 133 human skeletal samples from a wide geographical and temporal range, identified by the presence of skeletal lesions typical of chronic disseminated tuberculosis. Given the inherent challenges working with ancient mycobacteria, strict criteria must be used and primer/probe design continually re-evaluated as new data from bacteria become available. Seven samples tested positive for multiple MTBC loci, supporting them as strong candidates for downstream analyses. Using strict and conservative criteria, qPCR remains a fast and effective screening tool when compared with screening by more expensive sequencing and enrichment technologies.

Secrets in the bones Neanderthal Man In Search of Lost Genomes Svante Pääbo Basic Books, 2014. 285 pp.

![Figure][1] The Neanderthal is not entirely extinct. In fact, some part of this otherwise ancient hominin remains among us today, manifest in about 2% of the DNA of many modern humans as a result of ancient interbreeding between our species. One strand of Svante Paabo's Neanderthal Man explains the nearly 20 years of research that eventually led to this finding. Paabo provides a riveting, personal account of the development of paleogenetics and the technical revolution that made the field possible. The memoir also traces the sometimes unconventional path that led to his becoming a globe-trotting professor. ![Figure][1] Neanderthal type material PHOTO: SCIENCE PHOTO LIBRARY/CORBIS The author's success has stemmed from his determination to pursue what many assumed were unknowable answers to fundamental questions about human origins. In a career replete with unexpected twists, Paabo conducted secret experiments on Egyptian mummies, researched modern kangaroo rats as well as ancient cave bears, and even used genetic analyses of his own students' sperm to advance his research. His journey was certainly not without its difficulties, and he writes about complications with merciless candor. For instance, he leaves out few raw details when discussing his own insecurities, shortcomings of his and others' research, political and cultural obstacles that often impede scientific progress, and the affairs (romantic as well as scientific) that at times threatened to turn research collaborators into competitors. The author effectively conveys both the broad implications of his work and his concern for scientific details. While reading, you may find yourself with the sudden urge to wash your hands when Paabo expounds on the ubiquitous nature of DNA contamination and the absolute necessity for a completely sterile workspace. These fears were particularly salient in the earlier years, when he required that all data of any importance be independently replicated by scientists in other laboratories. For perhaps his most celebrated finding—that there exists a “clearly discernible genetic contribution from Neanderthals to people outside Africa”—Paabo would settle for no fewer than three independent lines of statistical evidence before publishing. To nonscientists, this self-professed meticulousness may at times seem to border on neurotic. But in a time of frequent headlines about scientific fraud, his portrayal of the internal and external struggles that accompany a career in academia refreshingly illustrates the high standards required for a rigorous research program. At its core, the book is about exposing secrets: from the nucleotide sequences laboriously pried from 40,000-year-old bones to the inner workings of a prominent researcher's mind. Whether unacquainted with Paabo's work or regular followers of his publications, readers will find that Neanderthal Man provides a nonpareil account of the development of the field of ancient DNA. [1]: pending:yes

Neanderthal man: in search of lost genomes

What can we learn from the genomes of our closest evolutionary relatives? Neanderthal Man tells the story of geneticist Svante Paabo's mission to answer this question, and recounts his ultimately successful efforts to genetically define what makes us different from our Neanderthal cousins. Beginning with the study of DNA in Egyptian mummies in the early 1980s and culminating in the sequencing of the Neanderthal genome in 2010, Neanderthal Man describes the events, intrigues, failures, and triumphs of these scientifically rich years through the lens of the pioneer and inventor of the field of ancient DNA. We learn that Neanderthal genes offer a unique window into the lives of our hominin relatives and may hold the key to unlocking the mystery of why humans survived while Neanderthals went extinct. Drawing on genetic and fossil clues, Paabo explores what is known about the origin of modern humans and their relationship to the Neanderthals and describes the fierce debate surrounding the nature of the two species' interactions. His findings have not only redrawn our family tree, but recast the fundamentals of human history--the biological beginnings of fully modern Homo sapiens, the direct ancestors of all people alive today. A riveting story about a visionary researcher and the nature of scientific inquiry, Neanderthal Man offers rich insight into the fundamental question of who we are.

Evolution: The human puzzle

Henry Gee relishes the memoir of Svante Pääbo, a leader in the field of ancient DNA.

A Paleogenomic Perspective on Evolution and Gene Function: New Insights from Ancient DNA

The publication of partial and complete paleogenomes within the last few years has reinvigorated research in ancient DNA. No longer limited to short fragments of mitochondrial DNA, inference of evolutionary processes through time can now be investigated from genome-wide data sampled as far back as 700,000 years. Tremendous insights have been made, in particular regarding the hominin lineage. With rare exception, however, a paleogenomic perspective has been mired by the quality and quantity of recoverable DNA. Though conceptually simple, extracting ancient DNA remains challenging, and sequencing ancient genomes to high coverage remains prohibitively expensive for most laboratories. Still, with improvements in DNA isolation and declining sequencing costs, the taxonomic and geographic purview of paleogenomics is expanding at a rapid pace. With improved capacity to screen large numbers of samples for those with high proportions of endogenous ancient DNA, paleogenomics is poised to become a key technology to better understand recent evolutionary events.

Carles Lalueza-Fox

Carles Lalueza-Fox

Ancient DNA studies: new perspectives on old samples

In spite of past controversies, the field of ancient DNA is now a reliable research area due to recent methodological improvements. A series of recent large-scale studies have revealed the true potential of ancient DNA samples to study the processes of evolution and to test models and assumptions commonly used to reconstruct patterns of evolution and to analyze population genetics and palaeoecological changes. Recent advances in DNA technologies, such as next-generation sequencing make it possible to recover DNA information from archaeological and paleontological remains allowing us to go back in time and study the genetic relationships between extinct organisms and their contemporary relatives. With the next-generation sequencing methodologies, DNA sequences can be retrieved even from samples (for example human remains) for which the technical pitfalls of classical methodologies required stringent criteria to guaranty the reliability of the results. In this paper, we review the methodologies applied to ancient DNA analysis and the perspectives that next-generation sequencing applications provide in this field.

Resurrecting ancient animal genomes: The extinct moa and more

Recently two developments have had a major impact on the field of ancient DNA (aDNA). First, new advances in DNA sequencing, in combination with improved capture/enrichment methods, have resulted in the recovery of orders of magnitude more DNA sequence data from ancient animals. Second, there has been an increase in the range of tissue types employed in aDNA. Hair in particular has proven to be very successful as a source of DNA because of its low levels of contamination and high level of ancient endogenous DNA. These developments have resulted in significant advances in our understanding of recently extinct animals: namely their evolutionary relationships, physiology, and even behaviour. Hair has been used to recover the first complete ancient nuclear genome, that of the extinct woolly mammoth, which then facilitated the expression and functional analysis of haemoglobins. Finally, we speculate on the consequences of these developments for the possibility of recreating extinct animals.

Application and comparison of large-scale solution-based DNA capture-enrichment methods on ancient DNA

The development of second-generation sequencing technologies has greatly benefitted the field of ancient DNA (aDNA). Its application can be further exploited by the use of targeted capture-enrichment methods to overcome restrictions posed by low endogenous and contaminating DNA in ancient samples. We tested the performance of Agilent's SureSelect and Mycroarray's MySelect in-solution capture systems on Illumina sequencing libraries built from ancient maize to identify key factors influencing aDNA capture experiments. High levels of clonality as well as the presence of multiple-copy sequences in the capture targets led to biases in the data regardless of the capture method. Neither method consistently outperformed the other in terms of average target enrichment, and no obvious difference was observed either when two tiling designs were compared. In addition to demonstrating the plausibility of capturing aDNA from ancient plant material, our results also enable us to provide useful recommendations for those planning targeted-sequencing on aDNA.

True single-molecule DNA sequencing of a pleistocene horse bone

Second-generation sequencing platforms have revolutionized the field of ancient DNA, opening access to complete genomes of past individuals and extinct species. However, these platforms are dependent on library construction and amplification steps that may result in sequences that do not reflect the original DNA template composition. This is particularly true for ancient DNA, where templates have undergone extensive damage post-mortem. Here, we report the results of the first "true single molecule sequencing" of ancient DNA. We generated 115.9 Mb and 76.9 Mb of DNA sequences from a permafrost-preserved Pleistocene horse bone using the Helicos HeliScope and Illumina GAIIx platforms, respectively. We find that the percentage of endogenous DNA sequences derived from the horse is higher among the Helicos data than Illumina data. This result indicates that the molecular biology tools used to generate sequencing libraries of ancient DNA molecules, as required for second-generation sequencing, introduce biases into the data that reduce the efficiency of the sequencing process and limit our ability to fully explore the molecular complexity of ancient DNA extracts. We demonstrate that simple modifications to the standard Helicos DNA template preparation protocol further increase the proportion of horse DNA for this sample by threefold. Comparison of Helicos-specific biases and sequence errors in modern DNA with those in ancient DNA also reveals extensive cytosine deamination damage at the 3' ends of ancient templates, indicating the presence of 3'-sequence overhangs. Our results suggest that paleogenomes could be sequenced in an unprecedented manner by combining current second- and third-generation sequencing approaches.

Moa's Ark or volant ghosts of Gondwana? Insights from nineteen years of ancient DNA research on the extinct moa (Aves: Dinornithiformes) of New Zealand

The moa (Aves: Dinornithiformes) of New Zealand represent one of the extinct iconic taxa that define the field of ancient DNA (aDNA), and after almost two decades of genetic scrutiny of bones, feathers, coprolites, mummified tissue, eggshell, and sediments, our knowledge of these prehistoric giants has increased significantly. Thanks to molecular and morphological-based research, the insights that have been obtained into moa phylogenetics, phylogeography, and palaeobiology exceeds that of any other extinct taxon. This review documents the strengths of applying a multidisciplinary approach when studying extinct taxa but also shows that cross-disciplinary controversies still remain at the most fundamental levels, with highly conflicting interpretations derived from aDNA and morphology. Moa species diversity, for example, is still heavily debated, as well as their relationship with other ratites and the mode of radiation. In addition to increasing our knowledge on a lineage of extinct birds, further insights into these aspects can clarify some of the basal splits in avian evolution, and the evolutionary implications of the breakup of the prehistoric supercontinent Gondwana. Did a flightless moa ancestor drift away on proto New Zealand (Moa's Ark) or did a volant ancestor arrive by flight? Here we provide an overview of 19 years of aDNA research on moa, critically assess the attempts and controversies in placing the moa lineage among palaeognath birds, and discuss the factors that facilitated the extensive radiation of moa. Finally, we identify the most obvious gaps in the current knowledge to address the future potential research areas in moa genetics.

Profiling the Dead: Generating Microsatellite Data from Fossil Bones of Extinct Megafauna—Protocols, Problems, and Prospects

We present the first set of microsatellite markers developed exclusively for an extinct taxon. Microsatellite data have been analysed in thousands of genetic studies on extant species but the technology can be problematic when applied to low copy number (LCN) DNA. It is therefore rarely used on substrates more than a few decades old. Now, with the primers and protocols presented here, microsatellite markers are available to study the extinct New Zealand moa (Aves: Dinornithiformes) and, as with single nucleotide polymorphism (SNP) technology, the markers represent a means by which the field of ancient DNA can (preservation allowing) move on from its reliance on mitochondrial DNA. Candidate markers were identified using high throughput sequencing technology (GS-FLX) on DNA extracted from fossil moa bone and eggshell. From the ‘shotgun’ reads, >60 primer pairs were designed and tested on DNA from bones of the South Island giant moa (Dinornis robustus). Six polymorphic loci were characterised and used to assess measures of genetic diversity. Because of low template numbers, typical of ancient DNA, allelic dropout was observed in 36–70% of the PCR reactions at each microsatellite marker. However, a comprehensive survey of allelic dropout, combined with supporting quantitative PCR data, allowed us to establish a set of criteria that maximised data fidelity. Finally, we demonstrated the viability of the primers and the protocols, by compiling a full Dinornis microsatellite dataset representing fossils of c. 600–5000 years of age. A multi-locus genotype was obtained from 74 individuals (84% success rate), and the data showed no signs of being compromised by allelic dropout. The methodology presented here provides a framework by which to generate and evaluate microsatellite data from samples of much greater antiquity than attempted before, and opens new opportunities for ancient DNA research.

Validation of the MiniFiler™ Kit in archaeological samples

We evaluate the usefulness of MiniFiler™ Kit in the field of ancient DNA. A set of samples belonging to different locations from Iberian Peninsula, with ages ranging from Neolithic to XVII century, was tested. Results could be replicated in only one burial site, probably due to the taphonomic conditions. Other cases could only produce partial or none genetic profiles.

Identification of Microsatellites from An Extinct Moa Species Using High-Throughput (454) Sequence Data

Genetic variation in microsatellites is rarely examined in the field of ancient DNA (aDNA) due to the low quantity of nuclear DNA in the fossil record together with the lack of characterized nuclear markers in extinct species. 454 sequencing platforms provide a new high-throughput technology capable of generating up to 1 gigabases per run as short (200-400-bp) read lengths. 454 data were generated from the fossil bone of an extinct New Zealand moa (Aves: Dinornithiformes). We identified numerous short tandem repeat (STR) motifs, and here present the successful isolation and characterization of one polymorphic microsatellite (Moa_MS2). Primers designed to flank this locus amplified all three moa species tested here. The presented method proved to be a fast and efficient way of identifying microsatellite markers in ancient DNA templates and, depending on biomolecule preservation, has the potential of enabling high-resolution population genetic studies of extinct taxa. As sequence read lengths of the 454 platforms and its competitors (e.g., the SOLEXA and SOLiD platforms) increase, this approach will become increasingly powerful in identifying microsatellites in extinct (and extant) organisms, and will afford new opportunities to study past biodiversity and extinction processes.

Wanted: Clean Neandertal DNA

It usually pays to have more than one team working on a scientific problem. Take the early work on sequencing Neandertal nuclear DNA, for example. In 2005, Svante Pääbo and his crew at the Max Planck Institute for Evolutionary Anthropology in Leipzig, Germany, gave Neandertal DNA samples to Edward Rubin's group at the U.S. Department of Energy Joint Genome Institute (JGI) in Walnut Creek, California. Together, the two groups helped revitalize the field of ancient DNA by producing more than a million bases of Neandertal nuclear genome sequence (Science, 17 November 2006, p. 1068). Equally important: Differences in their results alerted the field to what later proved to be contaminating DNA in Pääbo's analysis.

The Effect of Ancient DNA Damage on Inferences of Demographic Histories

The field of ancient DNA (aDNA) is casting new light on many evolutionary questions. However, problems associated with the postmortem instability of DNA may complicate the interpretation of aDNA data. For example, in population genetic studies, the inclusion of damaged DNA may inflate estimates of diversity. In this paper, we examine the effect of DNA damage on population genetic estimates of ancestral population size. We simulate data using standard coalescent simulations that include postmortem damage and show that estimates of effective population sizes are inflated around, or right after, the sampling time of the ancestral DNA sequences. This bias leads to estimates of increasing, and then decreasing, population sizes, as observed in several recently published studies. We reanalyze a recently published data set of DNA sequences from the Bison (Bison bison/Bison priscus) and show that the signal for a change in effective population size in this data set vanishes once the effects of putative damage are removed. Our results suggest that population genetic analyses of aDNA sequences, which do not accurately account for damage, should be interpreted with great caution.

The Dawn of Stone Age Genomics

PALEOGENETICSDNA from a 38,000-year-old Neandertal, including results reported on page [1113][1] of this issue of Science , is revitalizing the once-moribund field of ancient DNA, and it promises a fresh perspective on how we differ from our closest relatives. ([Read more][2].) [1]: http://www.sciencemag.org/cgi/content/short/314/5802/1113 [2]: http://www.sciencemag.org/cgi/content/full/314/5802/1068