Ancient Biomolecules and Evolutionary Inference.

Over the past three decades, studies of ancient biomolecules-particularly ancient DNA, proteins, and lipids-have revolutionized our understanding of evolutionary history. Though initially fraught with many challenges, today the field stands on firm foundations. Researchers now successfully retrieve nucleotide and amino acid sequences, as well as lipid signatures, from progressively older samples, originating from geographic areas and depositional environments that, until recently, were regarded as hostile to long-term preservation of biomolecules. Sampling frequencies and the spatial and temporal scope of studies have also increased markedly, and with them the size and quality of the data sets generated. This progress has been made possible by continuous technical innovations in analytical methods, enhanced criteria for the selection of ancient samples, integrated experimental methods, and advanced computational approaches. Here, we discuss the history and current state of ancient biomolecule research, its applications to evolutionary inference, and future directions for this young and exciting field.

[1]  Mattias Jakobsson,et al.  The genome of a Late Pleistocene human from a Clovis burial site in western Montana , 2014, Nature.

[2]  Janet Kelso,et al.  Neandertal and Denisovan DNA from Pleistocene sediments , 2017, Science.

[3]  Kirk C Hansen,et al.  Preserved Proteins from Extinct Bison latifrons Identified by Tandem Mass Spectrometry; Hydroxylysine Glycosides are a Common Feature of Ancient Collagen* , 2015, Molecular & Cellular Proteomics.

[4]  Ronald T Raines,et al.  Collagen structure and stability. , 2009, Annual review of biochemistry.

[5]  Michael Buckley,et al.  Identification of a new hominin bone from Denisova Cave, Siberia using collagen fingerprinting and mitochondrial DNA analysis , 2016, Scientific Reports.

[6]  Stefan Schouten,et al.  Distributional variations in marine crenarchaeotal membrane lipids: a new tool for reconstructing ancient sea water temperatures? , 2002 .

[7]  Keith Dobney,et al.  Genomic and archaeological evidence suggest a dual origin of domestic dogs , 2016, Science.

[8]  Eske Willerslev,et al.  Improving access to endogenous DNA in ancient bones and teeth , 2015, Scientific Reports.

[9]  Søren Brunak,et al.  Early Divergent Strains of Yersinia pestis in Eurasia 5,000 Years Ago , 2015, Cell.

[10]  James Haile,et al.  Ancient DNA reveals late survival of mammoth and horse in interior Alaska , 2009, Proceedings of the National Academy of Sciences.

[11]  Yong J. Kil,et al.  Byonic: Advanced Peptide and Protein Identification Software , 2012, Current protocols in bioinformatics.

[12]  J. Maguire,et al.  Solution Hybrid Selection with Ultra-long Oligonucleotides for Massively Parallel Targeted Sequencing , 2009, Nature Biotechnology.

[13]  Georg K. A. Hochberg,et al.  Reconstructing Ancient Proteins to Understand the Causes of Structure and Function. , 2017, Annual review of biophysics.

[14]  R. Evershed,et al.  Free and bound fatty acid oxidation products in archaeological ceramic vessels , 1998, Proceedings of the Royal Society of London. Series B: Biological Sciences.

[15]  Yong Wang,et al.  An Aboriginal Australian Genome Reveals Separate Human Dispersals into Asia , 2011, Science.

[16]  David M. Rocke,et al.  Demonstration of Protein-Based Human Identification Using the Hair Shaft Proteome , 2016, PloS one.

[17]  M. Mann,et al.  MaxQuant enables high peptide identification rates, individualized p.p.b.-range mass accuracies and proteome-wide protein quantification , 2008, Nature Biotechnology.

[18]  Philip L. F. Johnson,et al.  A Draft Sequence of the Neandertal Genome , 2010, Science.

[19]  M. Hofreiter,et al.  A mitogenomic timetree for Darwin’s enigmatic South American mammal Macrauchenia patachonica , 2017, Nature Communications.

[20]  M. Thomas P. Gilbert,et al.  Resolution of the type material of the Asian elephant, Elephas maximus Linnaeus, 1758 (Proboscidea, Elephantidae) , 2014 .

[21]  W Wackernagel,et al.  Adsorption of plasmid DNA to mineral surfaces and protection against DNase I , 1991, Applied and environmental microbiology.

[22]  Stephan C. Schuster,et al.  Response to Comment on "Whole-Genome Shotgun Sequencing of Mitochondria from Ancient Hair Shafts" , 2008, Science.

[23]  C. Wiuf,et al.  Statistical evidence for miscoding lesions in ancient DNA templates. , 2001, Molecular biology and evolution.

[24]  Rob Knight,et al.  Insights from Characterizing Extinct Human Gut Microbiomes , 2012, PloS one.

[25]  Richard P. Evershed,et al.  QUANTIFICATION AND DISTRIBUTION OF LIPID IN ARCHAEOLOGICAL CERAMICS: IMPLICATIONS FOR SAMPLING POTSHERDS FOR ORGANIC RESIDUE ANALYSIS AND THE CLASSIFICATION OF VESSEL USE , 1993 .

[26]  P. Kapranov,et al.  True single-molecule DNA sequencing of a pleistocene horse bone. , 2011, Genome research.

[27]  Michael Buckley,et al.  A Molecular Phylogeny of Plesiorycteropus Reassigns the Extinct Mammalian Order ‘Bibymalagasia’ , 2013, PloS one.

[28]  Lee E Kirsch,et al.  The estimation of glutaminyl deamidation and aspartyl cleavage rates in glucagon. , 2004, International journal of pharmaceutics.

[29]  Michael Buckley,et al.  Species identification by analysis of bone collagen using matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry. , 2009, Rapid communications in mass spectrometry : RCM.

[30]  James Haile,et al.  Species-specific responses of Late Quaternary megafauna to climate and humans , 2011, Nature.

[31]  Eleanor J. Green,et al.  Novel Substrates as Sources of Ancient DNA: Prospects and Hurdles , 2017, Genes.

[32]  Oliver Smith,et al.  A new model for ancient DNA decay based on paleogenomic meta-analysis , 2017 .

[33]  Janet Kelso,et al.  Palaeoproteomic evidence identifies archaic hominins associated with the Châtelperronian at the Grotte du Renne , 2016, Proceedings of the National Academy of Sciences.

[34]  Richard J. Lavallee,et al.  Optimized fast and sensitive acquisition methods for shotgun proteomics on a quadrupole orbitrap mass spectrometer. , 2012, Journal of proteome research.

[35]  R Buick,et al.  Archean molecular fossils and the early rise of eukaryotes. , 1999, Science.

[36]  Charly Favier,et al.  Five thousand years of tropical lake sediment DNA records from Benin , 2017 .

[37]  M. Jakobsson,et al.  Separating endogenous ancient DNA from modern day contamination in a Siberian Neandertal , 2014, Proceedings of the National Academy of Sciences.

[38]  Milda Pucetaite,et al.  Molecular signatures of fossil leaves provide unexpected new evidence for extinct plant relationships , 2017, Nature Ecology & Evolution.

[39]  Kendra Sirak,et al.  Optimal Ancient DNA Yields from the Inner Ear Part of the Human Petrous Bone , 2015, PloS one.

[40]  D. Reich,et al.  Population Structure and Eigenanalysis , 2006, PLoS genetics.

[41]  Keith Dobney,et al.  Using ancient DNA to study the origins and dispersal of ancestral Polynesian chickens across the Pacific , 2014, Proceedings of the National Academy of Sciences.

[42]  Richard P. Evershed,et al.  Resolving the bulk δ15N values of ancient human and animal bone collagen via compound-specific nitrogen isotope analysis of constituent amino acids , 2010 .

[43]  Maria Perla Colombini,et al.  Organic Mass Spectrometry in Art and Archaeology , 2009 .

[44]  Mark Horton,et al.  A novel marine dietary indicator utilising compound-specific bone collagen amino acid δ13C values of ancient humans , 2005 .

[45]  A. Pearson Lipidomics for Geochemistry , 2014 .

[46]  Jesper V Olsen,et al.  Pathogens and host immunity in the ancient human oral cavity , 2014, Nature Genetics.

[47]  E. Willerslev,et al.  More on contamination: the use of asymmetric molecular behavior to identify authentic ancient human DNA. , 2007, Molecular biology and evolution.

[48]  Søren Brunak,et al.  Population genomics of Bronze Age Eurasia , 2015, Nature.

[49]  Charlotte L. Oskam,et al.  The half-life of DNA in bone: measuring decay kinetics in 158 dated fossils , 2012, Proceedings of the Royal Society B: Biological Sciences.

[50]  Derek E. G. Briggs,et al.  The biomolecular paleontology of continental fossils , 2000, Paleobiology.

[51]  Richard P. Evershed,et al.  Earliest evidence for cheese making in the sixth millennium bc in northern Europe , 2012, Nature.

[52]  Richard P. Evershed,et al.  Experimental approaches to the interpretation of absorbed organic residues in archaeological ceramics , 2008 .

[53]  W. Koh,et al.  Single-cell genome sequencing: current state of the science , 2016, Nature Reviews Genetics.

[54]  S. Pääbo The Human Condition—A Molecular Approach , 2014, Cell.

[55]  Roman Fischer,et al.  Animal origin of 13th-century uterine vellum revealed using noninvasive peptide fingerprinting , 2015, Proceedings of the National Academy of Sciences.

[56]  P. Donnelly,et al.  Inference of population structure using multilocus genotype data. , 2000, Genetics.

[57]  Daniel J. Condon,et al.  Fossil steroids record the appearance of Demospongiae during the Cryogenian period , 2009, Nature.

[58]  R. Evershed,et al.  The significance of petroleum bitumen in ancient Egyptian mummies , 2016, Philosophical Transactions of the Royal Society A: Mathematical, Physical and Engineering Sciences.

[59]  Aurélien Ginolhac,et al.  A comparative study of ancient environmental DNA to pollen and macrofossils from lake sediments reveals taxonomic overlap and additional plant taxa , 2013 .

[60]  Thomas Levy,et al.  Earliest date for milk use in the Near East and southeastern Europe linked to cattle herding , 2008, Nature.

[61]  A. B. Robinson,et al.  Deamidation of glutaminyl and asparaginyl residues in peptides and proteins. , 1974, Current topics in cellular regulation.

[62]  Clément Zanolli,et al.  Neanderthal and Denisova tooth protein variants in present-day humans , 2017, PloS one.

[63]  Marie Besse,et al.  The Beaker Phenomenon and the Genomic Transformation of Northwest Europe , 2018, Nature.

[64]  M. Slatkin,et al.  Joint Estimation of Contamination, Error and Demography for Nuclear DNA from Ancient Humans , 2015, bioRxiv.

[65]  Beth Shapiro,et al.  Rise and Fall of the Beringian Steppe Bison , 2004, Science.

[66]  Enilson Antonio Sallum,et al.  Proteomic analysis of human dental cementum and alveolar bone. , 2013, Journal of proteomics.

[67]  Mattias Jakobsson,et al.  Tracing the peopling of the world through genomics , 2017, Nature.

[68]  Michael Schall,et al.  New strategies for characterizing ancient proteins using matrix-assisted laser desorption ionization mass spectrometry , 2000 .

[69]  Ming Li,et al.  PEAKS: powerful software for peptide de novo sequencing by tandem mass spectrometry. , 2003, Rapid communications in mass spectrometry : RCM.

[70]  Mark Horton,et al.  Volatile Compounds in Archaeological Plant Remains and the Maillard Reaction During Decay of Organic Matter , 1997 .

[71]  T. Devièse,et al.  RECONSTRUCTING ANCIENT YEMENI COMMERCIAL ROUTES DURING THE MIDDLE AGES USING STRUCTURAL CHARACTERIZATION OF TERPENOID RESINS , 2008 .

[72]  Marie Soressi,et al.  Site-specific deamidation of glutamine: a new marker of bone collagen deterioration. , 2012, Rapid communications in mass spectrometry : RCM.

[73]  Roger E. Summons,et al.  2-Methylhopanoids as biomarkers for cyanobacterial oxygenic photosynthesis , 1999, Nature.

[74]  Anita Quye,et al.  Characterisation of "bog butter" using a combination of molecular and isotopic techniques. , 2004, The Analyst.

[75]  N. A. Stewart,et al.  Sex determination of human remains from peptides in tooth enamel , 2017, Proceedings of the National Academy of Sciences.

[76]  Deepak Vashishth,et al.  Peptide sequences from the first Castoroides ohioensis skull and the utility of old museum collections for palaeoproteomics , 2016, Proceedings of the Royal Society B: Biological Sciences.

[77]  Yang Wang,et al.  Expansion of C4 ecosystems as an indicator of global ecological change in the late Miocene , 1993, Nature.

[78]  A. von Haeseler,et al.  DNA sequences from multiple amplifications reveal artifacts induced by cytosine deamination in ancient DNA. , 2001, Nucleic acids research.

[79]  Janet Kelso,et al.  Nuclear DNA sequences from the Middle Pleistocene Sima de los Huesos hominins , 2016, Nature.

[80]  Samuel T. Turvey,et al.  Ancient proteins resolve the evolutionary history of Darwin’s South American ungulates , 2015, Nature.

[81]  Daniel H. Huson,et al.  Neanderthal behaviour, diet, and disease inferred from ancient DNA in dental calculus , 2017, Nature.

[82]  D. Reich,et al.  Genome-wide patterns of selection in 230 ancient Eurasians , 2015, Nature.

[83]  John A. Kamm,et al.  Terminal Pleistocene Alaskan genome reveals first founding population of Native Americans , 2018, Nature.

[84]  M. Jakobsson,et al.  Origins and Genetic Legacy of Neolithic Farmers and Hunter-Gatherers in Europe , 2012, Science.

[85]  Elizabeth C. Theil,et al.  A role for iron and oxygen chemistry in preserving soft tissues, cells and molecules from deep time , 2014, Proceedings of the Royal Society B: Biological Sciences.

[86]  A. Tseng,et al.  Partial amino acid sequence of osteocalcin from an extinct species of ratite bird. , 1990, Biochemistry international.

[87]  Robin Bendrey,et al.  The Earliest Horse Harnessing and Milking , 2009, Science.

[88]  B. Ma,et al.  De Novo Sequencing and Homology Searching‡‡* , 2011, Molecular & Cellular Proteomics.

[89]  Asan,et al.  Altitude adaptation in Tibet caused by introgression of Denisovan-like DNA , 2014, Nature.

[90]  Stanley H. Ambrose,et al.  Probing dietary change of the Kwädąy Dän Ts'ìnchį individual, an ancient glacier body from British Columbia: I. Complementary use of marine lipid biomarker and carbon isotope signatures as novel indicators of a marine diet , 2008 .

[91]  Jaume Bertranpetit,et al.  Nuclear Gene Indicates Coat-Color Polymorphism in Mammoths , 2006, Science.

[92]  Philip L. F. Johnson,et al.  The complete genome sequence of a Neanderthal from the Altai Mountains , 2013 .

[93]  Keith Dobney,et al.  Sequencing ancient calcified dental plaque shows changes in oral microbiota with dietary shifts of the Neolithic and Industrial revolutions , 2013, Nature Genetics.

[94]  Timothy P. Cleland,et al.  Biologically and diagenetically derived peptide modifications in moa collagens , 2015, Proceedings of the Royal Society B: Biological Sciences.

[95]  Arne Ludwig,et al.  Experimental conditions improving in‐solution target enrichment for ancient DNA , 2017, Molecular ecology resources.

[96]  Diana J. Kao,et al.  Parallel adaptive radiations in two major clades of placental mammals , 2001, Nature.

[97]  Christian Rolando,et al.  Proteins in Art, Archaeology, and Paleontology: From Detection to Identification. , 2016, Chemical reviews.

[98]  R. Nielsen,et al.  Evidence for archaic adaptive introgression in humans , 2015, Nature Reviews Genetics.

[99]  M. Huber,et al.  Subtropical Arctic Ocean temperatures during the Palaeocene/Eocene thermal maximum , 2006, Nature.

[100]  M. Thomas P. Gilbert,et al.  mapDamage: testing for damage patterns in ancient DNA sequences , 2011, Bioinform..

[101]  Richard P. Evershed,et al.  High throughput screening of organic residues in archaeological potsherds using direct acidified methanol extraction , 2014 .

[102]  P. Taberlet,et al.  Fifty Thousand Years of Arctic Vegetation and Megafaunal Diet 1 Reconstruction of Arctic Vegetation from Permafrost Samples 121 , 2022 .

[103]  L. Excoffier,et al.  Robust Demographic Inference from Genomic and SNP Data , 2013, PLoS genetics.

[104]  M. Wilm,et al.  Analytical properties of the nanoelectrospray ion source. , 1996, Analytical chemistry.

[105]  Ivor Karavanić,et al.  The Genomic History of Southeastern Europe , 2017, Nature.

[106]  A. Krogh,et al.  Ancient human genome sequence of an extinct Palaeo-Eskimo , 2010, Nature.

[107]  Carl Heron,et al.  Analysis of organic residues of archaeological origin by high-temperature gas chromatography and gas chromatography-mass spectrometry , 1990 .

[108]  James Haile,et al.  Ancient Biomolecules from Deep Ice Cores Reveal a Forested Southern Greenland , 2007, Science.

[109]  Marshall Bern,et al.  Expansion for the Brachylophosaurus canadensis Collagen I Sequence and Additional Evidence of the Preservation of Cretaceous Protein. , 2017, Journal of proteome research.

[110]  Martin Sikora,et al.  Pulling out the 1%: whole-genome capture for the targeted enrichment of ancient DNA sequencing libraries. , 2013, American journal of human genetics.

[111]  Ludovic Antoine Alexandre,et al.  Improving ancient DNA read mapping against modern reference genomes , 2015 .

[112]  James Haile,et al.  Author response: Protein sequences bound to mineral surfaces persist into deep time , 2016 .

[113]  G. Weinstock,et al.  Direct selection of human genomic loci by microarray hybridization , 2007, Nature Methods.

[114]  Damian Szklarczyk,et al.  Proteomic analysis of a pleistocene mammoth femur reveals more than one hundred ancient bone proteins. , 2012, Journal of proteome research.

[115]  E. Cappellini,et al.  Optimization of DNA Recovery and Amplification from Non-Carbonized Archaeobotanical Remains , 2014, PloS one.

[116]  M. Mann,et al.  Andromeda: a peptide search engine integrated into the MaxQuant environment. , 2011, Journal of proteome research.

[117]  L. Orlando,et al.  Reconstructing ancient genomes and epigenomes , 2015, Nature Reviews Genetics.

[118]  Ross Barnett,et al.  A phenol-chloroform protocol for extracting DNA from ancient samples. , 2012, Methods in molecular biology.

[119]  James Haile,et al.  Ancient and modern environmental DNA , 2015, Philosophical Transactions of the Royal Society B: Biological Sciences.

[120]  S. Clarke,et al.  Deamidation, isomerization, and racemization at asparaginyl and aspartyl residues in peptides. Succinimide-linked reactions that contribute to protein degradation. , 1987, The Journal of biological chemistry.

[121]  Eske Willerslev,et al.  Postglacial viability and colonization in North America’s ice-free corridor , 2016, Nature.

[122]  J. A. Taylor,et al.  Sequence database searches via de novo peptide sequencing by tandem mass spectrometry. , 1997, Rapid communications in mass spectrometry : RCM.

[123]  Aurélien Ginolhac,et al.  Characterization of ancient and modern genomes by SNP detection and phylogenomic and metagenomic analysis using PALEOMIX , 2014, Nature Protocols.

[124]  Michael Buckley,et al.  Ancient collagen reveals evolutionary history of the endemic South American ‘ungulates’ , 2015, Proceedings of the Royal Society B: Biological Sciences.

[125]  Katherine H. Freeman,et al.  Molecular and isotopic records of C4 grassland expansion in the late miocene , 2001 .

[126]  Olivia Cheronet,et al.  A minimally-invasive method for sampling human petrous bones from the cranial base for ancient DNA analysis. , 2017, BioTechniques.

[127]  F. Welker Elucidation of cross-species proteomic effects in human and hominin bone proteome identification through a bioinformatics experiment , 2018, BMC Evolutionary Biology.

[128]  G. Fiorentino,et al.  A multidisciplinary study of archaeological grape seeds , 2009, Naturwissenschaften.

[129]  Richard P. Evershed,et al.  ORGANIC RESIDUE ANALYSIS IN ARCHAEOLOGY: THE ARCHAEOLOGICAL BIOMARKER REVOLUTION* , 2008 .

[130]  Ryan T Fellers,et al.  Advancing Top-down Analysis of the Human Proteome Using a Benchtop Quadrupole-Orbitrap Mass Spectrometer. , 2017, Journal of proteome research.

[131]  A. Wilson,et al.  DNA sequences from the quagga, an extinct member of the horse family , 1984, Nature.

[132]  R. Aebersold,et al.  Mass spectrometry in proteomics. , 2001, Chemical reviews.

[133]  Eric A. Gaucher,et al.  An experimental phylogeny to benchmark ancestral sequence reconstruction , 2016, Nature Communications.

[134]  Julie Dunne,et al.  First dairying in green Saharan Africa in the fifth millennium bc , 2012, Nature.

[135]  Salima Ikram,et al.  Organic chemistry of balms used in the preparation of pharaonic meat mummies , 2013, Proceedings of the National Academy of Sciences.

[136]  John M. Hayes,et al.  Isotope-ratio-monitoring gas chromatography-mass spectrometry , 1978 .

[137]  Stephen Killops,et al.  An introduction to organic geochemistry , 1993 .

[138]  D. Minnikin,et al.  Molecular Biomarkers for Ancient Tuberculosis , 2012 .

[139]  Keith Dobney,et al.  Ancient DNA, pig domestication, and the spread of the Neolithic into Europe , 2007, Proceedings of the National Academy of Sciences.

[140]  Julie Dunne,et al.  Earliest direct evidence of plant processing in prehistoric Saharan pottery , 2016, Nature Plants.

[141]  R. Mägi,et al.  Upper Palaeolithic Siberian genome reveals dual ancestry of Native Americans , 2013, Nature.

[142]  Eric Coissac,et al.  Bioinformatic challenges for DNA metabarcoding of plants and animals , 2012, Molecular ecology.

[143]  Carsten Wiuf,et al.  Diverse Plant and Animal Genetic Records from Holocene and Pleistocene Sediments , 2003, Science.

[144]  Philip L. F. Johnson,et al.  Recalibrating Equus evolution using the genome sequence of an early Middle Pleistocene horse , 2013, Nature.

[145]  Swapan Mallick,et al.  Massive migration from the steppe was a source for Indo-European languages in Europe , 2015, Nature.

[146]  Keith Bennett,et al.  DNA from pollen: principles and potential , 2006 .

[147]  Julie Dunne,et al.  Widespread exploitation of the honeybee by early Neolithic farmers , 2016, Nature.

[148]  W. Michaelis,et al.  Structural investigations of sulphur-rich macromolecular oil fractions and a kerogen by sequential chemical degradation , 1992 .

[149]  Sabine Gaudzinski-Windheuser,et al.  Middle Pleistocene protein sequences from the rhinoceros genus Stephanorhinus and the phylogeny of extant and extinct Middle/Late Pleistocene Rhinocerotidae , 2017, PeerJ.

[150]  R. Durbin,et al.  Inference of human population history from individual whole-genome sequences. , 2011, Nature.

[151]  Peter C. Jordan,et al.  Earliest evidence for the use of pottery , 2013, Nature.

[152]  G. Eglinton,et al.  Molecular stratigraphy: a new tool for climatic assessment , 1986, Nature.

[153]  Matthias Meyer,et al.  A draft genome of Yersinia pestis from victims of the Black Death , 2011, Nature.

[154]  Philip L. F. Johnson,et al.  Targeted Investigation of the Neandertal Genome by Array-Based Sequence Capture , 2010, Science.

[155]  Anna J. Mukherjee,et al.  Direct chemical evidence for widespread dairying in prehistoric Britain , 2003, Proceedings of the National Academy of Sciences of the United States of America.

[156]  J. Lowenstein,et al.  Immunological reactions from fossil material. , 1981, Philosophical transactions of the Royal Society of London. Series B, Biological sciences.

[157]  Konrad A Hughen,et al.  Abrupt warming events drove Late Pleistocene Holarctic megafaunal turnover , 2015, Science.

[158]  Søren Brunak,et al.  Investigation of Human Cancers for Retrovirus by Low-Stringency Target Enrichment and High-Throughput Sequencing , 2015, Scientific Reports.

[159]  Bernhard Kuster,et al.  DMSO enhances electrospray response, boosting sensitivity of proteomic experiments , 2013, Nature Methods.

[160]  Roger E. Summons,et al.  Petroleum geology and geochemistry of the Middle Proterozoic McArthur Basin, Northern Australia: III. Composition of extractable hydrocarbons , 1988 .

[161]  J. R. Maxwell,et al.  Chemical Fossils: The Geological Fate of Steroids , 1982, Science.

[162]  Anders Albrechtsen,et al.  ANGSD: Analysis of Next Generation Sequencing Data , 2014, BMC Bioinformatics.

[163]  Leif Andersson,et al.  Prehistoric genomes reveal the genetic foundation and cost of horse domestication , 2014, Proceedings of the National Academy of Sciences.

[164]  Arne Ludwig,et al.  Ancient genomes revisit the ancestry of domestic and Przewalski’s horses , 2018, Science.

[165]  Baozhen Shan,et al.  De novo peptide sequencing by deep learning , 2017, Proceedings of the National Academy of Sciences.

[166]  T. Cleland,et al.  Solid Digestion of Demineralized Bone as a Method To Access Potentially Insoluble Proteins and Post-Translational Modifications. , 2018, Journal of proteome research.

[167]  W. Miller,et al.  Polar and brown bear genomes reveal ancient admixture and demographic footprints of past climate change , 2012, Proceedings of the National Academy of Sciences.

[168]  M. Thomas P. Gilbert,et al.  Application and comparison of large-scale solution-based DNA capture-enrichment methods on ancient DNA , 2011, Scientific reports.

[169]  J. Taubenberger,et al.  Integrating historical, clinical and molecular genetic data in order to explain the origin and virulence of the 1918 Spanish influenza virus. , 2001, Philosophical transactions of the Royal Society of London. Series B, Biological sciences.

[170]  Arne Ludwig,et al.  Ancient genomic changes associated with domestication of the horse , 2017, Science.

[171]  Evershed,et al.  Direct demonstration of milk as an element of archaeological economies , 1998, Science.

[172]  T. Lindahl Instability and decay of the primary structure of DNA , 1993, Nature.

[173]  Irving A. Breger,et al.  Geochemistry of lipids , 1966 .

[174]  N. Kelleher,et al.  Progress in Top-Down Proteomics and the Analysis of Proteoforms. , 2016, Annual review of analytical chemistry.