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LETTERdoi:10.1038/nature12960
Derived immune and ancestral pigmentation allelesin a 7,000-year-old Mesolithic EuropeanInigo Olalde1*, Morten E. Allentoft2*, Federico Sanchez-Quinto1, Gabriel Santpere1, Charleston W. K. Chiang3,Michael DeGiorgio4,5, Javier Prado-Martinez1, Juan Antonio Rodrıguez1, Simon Rasmussen6, Javier Quilez1, Oscar Ramırez1,Urko M. Marigorta1, Marcos Fernandez-Callejo1, Marıa Encina Prada7, Julio Manuel Vidal Encinas8, Rasmus Nielsen9,Mihai G. Netea10, John Novembre11, Richard A. Sturm12, Pardis Sabeti13,14, Tomas Marques-Bonet1,15, Arcadi Navarro1,15,16,17,Eske Willerslev2 & Carles Lalueza-Fox1
Ancient genomic sequences have started to reveal the origin and thedemographic impact of farmers from the Neolithic period spread-ing into Europe1–3. The adoption of farming, stock breeding andsedentary societies during the Neolithic may have resulted in adapt-ive changes in genes associated with immunity and diet4. However,the limited data available from earlier hunter-gatherers precludean understanding of the selective processes associated with this cru-cial transition to agriculture in recent human evolution. Here wesequence an approximately 7,000-year-old Mesolithic skeleton dis-covered at the La Brana-Arintero site in Leon, Spain, to retrieve acomplete pre-agricultural European human genome. Analysis ofthis genome in the context of other ancient samples suggests theexistence of a common ancient genomic signature across westernand central Eurasia from the Upper Paleolithic to the Mesolithic.The La Brana individual carries ancestral alleles in several skin pig-mentation genes, suggesting that the light skin of modern Europeanswas not yet ubiquitous in Mesolithic times. Moreover, we provideevidence that a significant number of derived, putatively adaptivevariants associated with pathogen resistance in modern Europeanswere already present in this hunter-gatherer.
Next-generation sequencing (NGS) technologies are revolution-izing the field of ancient DNA (aDNA), and have enabled the sequen-cing of complete ancient genomes5,6, such as that of Otzi, a Neolithichuman body found in the Alps1. However, very little is known of thegenetic composition of earlier hunter-gatherer populations from theMesolithic period (circa 10,000–5,000 years before present, BP; imme-diately preceding the Neolithic period).
The Iberian site called La Brana-Arintero was discovered in 2006when two male skeletons (named La Brana 1 and 2) were found in adeep cave system, 1,500 m above sea level in the Cantabrian mountainrange (Leon, Northwestern Spain) (Fig. 1a). The skeletons were datedto approximately 7,000 years BP (7,940–7,690 calibrated BP)7. Becauseof the cold environment and stable thermal conditions in the cave, thepreservation of these specimens proved to be exceptional (Fig. 1b). Weidentified a tooth from La Brana 1 with high human DNA content (48.4%)and sequenced this specimen to a final effective genomic depth-of-coverage of 3.403 (Extended Data Fig. 1).
We used several tests to assess the authenticity of the genome sequenceand to determine the amount of potential modern human contamina-tion. First, we observed that sequence reads from both the mitochondrial
*These authors contributed equally to this work.
1Institut de Biologia Evolutiva, CSIC-UPF, Barcelona 08003, Spain. 2Centre for GeoGenetics, Natural History Museum of Denmark, University of Copenhagen, DK-1350 Copenhagen K, Denmark.3Department of Ecology and Evolutionary Biology, University of California, Los Angeles, California 90095, USA. 4Department of Integrative Biology, University of California, Berkeley, California 94720, USA.5Department of Biology, Pennsylvania State University, 502 Wartik Laboratory, University Park, Pennsylvania 16802, USA. 6Center for Biological Sequence Analysis, Technical University of Denmark, DK-2800 Kongens Lyngby, Denmark. 7I.E.S.O. ’Los Salados’, Junta de Castilla y Leon, E-49600 Benavente, Spain. 8Junta de Castilla y Leon, Servicio de Cultura de Leon, E-24071 Leon, Spain. 9Center forTheoretical Evolutionary Genomics, University of California, Berkeley, California 94720, USA. 10Department of Medicine and Nijmegen Institute for Infection, Inflammation and Immunity, RadboudUniversity Nijmegen Medical Centre, 6500 Nijmegen, The Netherlands. 11Department of Human Genetics, University of Chicago, Illinois 60637, USA. 12Institute for Molecular Bioscience, MelanogenixGroup, The University of Queensland, Brisbane, Queensland 4072, Australia. 13Center for Systems Biology, Department of Organismic and Evolutionary Biology, Harvard University, Cambridge,Massachusetts 02138, USA. 14Broad Institute of the Massachusetts Institute of Technology and Harvard, Cambridge, Massachusetts 02142, USA. 15Institucio Catalana de Recerca i Estudis Avançats(ICREA), 08010 Barcelona, Catalonia, Spain. 16Centre de Regulacio Genomica (CRG), Barcelona 08003, Catalonia, Spain. 17National Institute for Bioinformatics (INB), Barcelona 08003, Catalonia, Spain.
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La Braña 1
Ötzi
Ajv70Ire8
Gok4
Ajv52
a b
c
La BraLa Braña-Arinteroa-Arinterositesite
La Braña-Arinterosite
Figure 1 | Geographic location and genetic affinities of the La Brana 1individual. a, Location of the La Brana-Arintero site (Spain). b, The La Brana 1skeleton as discovered in 2006. c, PCA based on the average of the Procrustestransformations of individual PCAs with La Brana 1 and each of the fiveNeolithic samples1,3. The reference populations are the Finnish HapMap,FINHM and POPRES. Population labels with labelling of ref. 12 with theaddition of FI (Finns) or LFI (late-settlement Finns). Ajv70, Ajv52, Ire8 andGok4 are Scandinavian Neolithic hunter-gatherers and a farmer, respectively3.Otzi is the Tyrolean Ice Man1.
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DNA (mtDNA) and the nuclear DNA of La Brana 1 showed the typicalancient DNA misincorporation patterns that arise from degradation ofDNA over time8 (Extended Data Fig. 2a, b). Second, we showed that theobserved number of human DNA fragments was negatively correlatedwith the fragment length (R2 . 0.92), as expected for ancient degradedDNA, and that the estimated rate of DNA decay was low and inagreement with predicted values9 (Extended Data Fig. 2c, d). We thenestimated the contamination rate in the mtDNA genome, assembled toa high depth-of-coverage (913), by checking for positions differingfrom the mtDNA genome (haplogroup U5b2c1) that was previouslyretrieved with a capture method2. We obtained an upper contamina-tion limit of 1.69% (0.75–2.6%, 95% confidence interval, CI) (Supplemen-tary Information). Finally, to generate a direct estimate of nuclear DNAcontamination, we screened for heterozygous positions (among readswith .43 coverage) in known polymorphic sites (Single NucleotidePolymorphism Database (dbSNP) build 137) at uniquely mapped sec-tions on the X chromosome6 (Supplementary Information). We foundthat the proportion of false heterozygous sites was 0.31%. Togetherthese results suggest low levels of contamination in the La Brana 1sequence data.
To investigate the relationship to extant European samples, we con-ducted a principal component analysis (PCA)10 and found that theapproximately 7,000-year-old Mesolithic sample was divergent fromextant European populations (Extended Data Fig. 3a, b), but was placedin proximity to northern Europeans (for example, samples from Swedenand Finland)11–14. Additional PCAs and allele-sharing analyses withancient Scandinavian specimens3 supported the genetic similarity ofthe La Brana 1 genome to Neolithic hunter-gatherers (Ajv70, Ajv52,Ire8) relative to Neolithic farmers (Gok4, Otzi) (Fig. 1c, Extended DataFigs 3c and 4). Thus, this Mesolithic individual from southwesternEurope represents a formerly widespread gene pool that seems to bepartially preserved in some modern-day northern European popula-tions, as suggested previously with limited genetic data2,3. We subse-quently explored the La Brana affinities to an ancient Upper Palaeolithicgenome from the Mal’ta site near Lake Baikal in Siberia15. Outgroup f3
and D statistics16,17, using different modern reference populations, sup-port that Mal’ta is significantly closer to La Brana 1 than to Asians ormodern Europeans (Extended Data Fig. 5 and Supplementary Infor-mation). These results suggest that despite the vast geographical dis-tance and temporal span, La Brana 1 and Mal’ta share common geneticancestry, indicating a genetic continuity in ancient western and centralEurasia. This observation matches findings of similar cultural artefactsacross time and space in Upper Paleolithic western Eurasia and Siberia,particularly the presence of anthropomorphic ‘Venus’ figurines thathave been recovered from several sites in Europe and Russia, includingthe Mal’ta site15. We also compared the genome-wide heterozygosity of
the La Brana 1 genome to a data set of modern humans with similarcoverage (3–43). The overall genomic heterozygosity was 0.042%,lower than the values observed in present day Asians (0.046–0.047%),Europeans (0.051–0.054%) and Africans (0.066–0.069%) (ExtendedData Fig. 6a). The effective population size, estimated from heterozyg-osity patterns, suggests a global reduction in population size of approxi-mately 20% relative to extant Europeans (Supplementary Information).Moreover, no evidence of tracts of autozygosity suggestive of inbreed-ing was observed (Extended Data Fig. 6b).
To investigate systematically the timing of selection events in therecent history of modern Europeans, we compared the La Brana gen-ome to modern populations at loci that have been categorized as ofinterest for their role in recent adaptive evolution. With respect to tworecent well-studied adaptations to changes in diet, we found the ancientgenome to carry the ancestral allele for lactose intolerance4 and approxi-mately five copies of the salivary amylase (AMY1) gene (Extended DataFig. 7 and Supplementary Information), a copy number compatiblewith a low-starch diet18. These results suggest the La Brana hunter-gatherer was poor at digesting milk and starch, supporting the hypo-theses that these abilities were selected for during the later transition toagriculture.
To expand the survey, we analysed a catalogue of candidate signalsfor recent positive selection based on whole-genome sequence vari-ation from the 1000 Genomes Project13, which included 35 candidatenon-synonymous variants, ten of which were detected uniquely in theCEU (Utah residents with northern and western European ancestry)sample 19. For each variant we assessed whether the Mesolithic genomecarried the ancestral or derived (putatively adaptive) allele.
Of the ten variants, the Mesolithic genome carried the ancestral andnon-selected allele as a homozygote in three regions: C12orf29 (a gene withunknown function), SLC45A2 (rs16891982) and SLC24A5 (rs1426654)(Table 1). The latter two variants are the two strongest known lociaffecting light skin pigmentation in Europeans20–22 and their ancestralalleles and associated haplotypes are either absent or segregate at verylow frequencies in extant Europeans (3% and 0% for SLC45A2 andSLC24A5, respectively) (Fig. 2). We subsequently examined all genesknown to be associated with pigmentation in Europeans22, and foundancestral alleles in MC1R, TYR and KITLG, and derived alleles inTYRP1, ASIP and IRF4 (Supplementary Information). Although theprecise phenotypic effects cannot currently be ascertained in a Europeangenetic background, results from functional experiments20 indicate thatthe allelic combination in this Mesolithic individual is likely to haveresulted in dark skin pigmentation and dark or brown hair. Furtherexamination revealed that this individual carried the HERC2 rs12913832*Csingle nucleotide polymorphism (SNP) and the associated homozygoushaplotype spanning the HERC2–OCA2 locus that is strongly associated
Table 1 | Mesolithic genome allelic state at 10 nonsynonymous variants recently selected in EuropeansAllelic state Gene Name SNP Amino-acid change Function
La Brana 1 carries thederived allele
PTX4 Pentraxin 4 rs2745098 Arg281Lys May be involved in innateimmunity
UHRF1BP1 UHRF1 binding protein 1 rs11755393 Gln454Arg Risk locus for systemiclupus erythematosus
GPATCH1 G patch domain containing 1 rs10421769 Leu520Ser Receptor for OmpA expressedby E. coli
WWOX WW domain-containing oxidoreductase rs12918952 Ala179Thr Acts as a tumour suppressor andhas a role in apoptosis
CCDC14 Coiled-coil domain-containing protein14
rs17310144 Thr365Pro Unknown
La Brana 1 carries boththe ancestral and the
derived allele
SETX Senataxin rs1056899 Val2587Ile Involved in spinocerebellar ataxiaand amyotrophic lateral sclerosis
TDRD12 Tudor domain containing 12 rs11881633 Glu413Lys UnknownLa Brana 1 retains the
ancestral alleleC12orf29 Chromosome 12 open reading frame 29 rs9262 Val238Leu Unknown
SLC45A2 Solute carrier family 45, member 2 rs16891982 Leu374Phe Associated with skin pigmentationSLC24A5 Solute carrier family 24, member 5 rs1426654 Ala111Thr Associated with skin pigmentation
RESEARCH LETTER
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with blue eye colour23. Moreover, a prediction of eye colour based ongenotypes at additional loci using HIrisPlex24 produced a 0.823 maximaland 0.672 minimal probability for being non-brown-eyed (Supplemen-tary Information). The genotypic combination leading to a predictedphenotype of dark skin and non-brown eyes is unique and no longerpresent in contemporary European populations. Our results indicatethat the adaptive spread of light skin pigmentation alleles was notcomplete in some European populations by the Mesolithic, and thatthe spread of alleles associated with light/blue eye colour may havepreceded changes in skin pigmentation.
For the remaining loci, La Brana 1 displayed the derived, putativelyadaptive variants in five cases, including three genes, PTX4, UHRF1BP1and GPATCH1 (ref. 19), involved in the immune system (Table 1 andExtended Data Fig. 8). GPATCH1 is associated with the risk of bacterialinfection. We subsequently determined the allelic states in 63 SNPsfrom 40 immunity genes with previous evidence for positive selectionand for carrying polymorphisms shown to influence susceptibility toinfections in modern Europeans (Supplementary Information). LaBrana 1 carries derived alleles in 24 genes (60%) that have a wide rangeof functions in the immune system: pattern recognition receptors,intracellular adaptor molecules, intracellular modulators, cytokinesand cytokine receptors, chemokines and chemokine receptors andeffector molecules. Interestingly, four out of six SNPs from the firstcategory are intracellular receptors of viral nucleic acids (TLR3, TLR8,IFIH1 (also known as MDA5) and LGP2)25.
Finally, to explore the functional regulation of the genome, we alsoassessed the La Brana 1 genotype at all expression quantitative trait loci(eQTL) regions associated to positive selection in Europeans (Sup-plementary Information). The most interesting finding is arguably thepredicted overexpression of eight immunity genes (36% of those with
described eQTLs), including three Toll-like receptor genes (TLR1, TLR2and TLR4) involved in pathogen recognition26.
These observations suggest that the Neolithic transition did not driveall cases of adaptive innovation on immunity genes found in modernEuropeans. Several of the derived haplotypes seen at high frequencytoday in extant Europeans were already present during the Mesolithic,as neutral standing variation or due to selection predating the Neolithic.De novo mutations that increased in frequency rapidly in response tozoonotic infections during the transition to farming should be iden-tified among those genes where La Brana 1 carries ancestral alleles.
To confirm whether the genomic traits seen at La Brana 1 can begeneralized to other Mesolithic populations, analyses of additional ancientgenomes from central and northern Europe will be needed. Nevertheless,this genome sequence provides the first insight as to how these hunter-gatherers are related to contemporary Europeans and other ancientpeoples in both Europe and Asia, and shows how ancient DNA can shedlight on the timing and nature of recent positive selection.
METHODS SUMMARYDNA was extracted from the La Brana 1 tooth specimen with a previously pub-lished protocol2. Indexed libraries were built from the ancient extract and sequencedon the Illumina HiSeq platform. Reads generated were mapped with BWA27 to thehuman reference genome (NCBI 37, hg19) after primer trimming. A metagenomicanalysis and taxonomic identification was generated with the remaining readsusing BLAST 2.2.271 and MEGAN4 (ref. 28) (Extended Data Fig. 9). SNP callingwas undertaken using a specific bioinformatic pipeline designed to account forancient DNA errors. Specifically, the quality of misincorporations likely caused byancient DNA damage was rescaled using the mapDamage2.0 software29, and a setof variants with a minimum read depth of 4 was produced with GATK30. Analysesincluding PCA10, Outgroup f3
16 and D statistics17 were performed to determine thepopulation affinities of this Mesolithic individual (Supplementary Information).
rs75
5678
77rs
1920
9718
8rs
3540
2rs
7937
1483
rs18
7050
917
rs73
0748
91rs
3540
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8763
63rs
8006
4515
rs10
4619
28rs
1119
1288
6rs
1118
4507
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7727
8088
rs14
0309
535
rs35
405
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3214
512
rs35
406
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0707
453
rs78
8550
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3540
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5795
9046
rs55
7466
63rs
1147
9453
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998
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6789
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1006
7723
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5630
314
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1633
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9142
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3400
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3539
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7850
5258
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8919
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1138
9109
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1142
3718
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1380
0566
1rs
1364
038
rs18
5145
rs14
9480
117
rs18
5146
rs73
0771
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7534
4149
rs25
0417
rs11
6828
782
rs11
3928
135
rs22
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1309
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1820
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6125
987
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393
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043
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6403
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2294
rs10
1087
2rs
2877
7rs
7736
6462
rs11
4048
710
rs75
0924
13rs
1890
0233
4rs
7307
7127
rs14
9427
409
rs19
0934
666
rs14
1821
415
rs80
2925
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1116
1849
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4525
rs74
4425
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2156
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rs75
6267
86rs
7580
4106
rs14
3538
859
rs18
5587
149
rs11
6688
269
rs22
4465
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1502
7178
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1016
2789
rs79
8754
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5572
8404
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7534
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8925
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5868
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1337
581
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6348
543
rs19
1207
217
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3165
868
rs18
8836
177
rs24
3335
4rs
2459
391
rs77
5757
93rs
1917
4093
4rs
1904
3625
4rs
7729
4837
rs14
2736
532
rs13
9737
156
rs26
7534
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1842
0002
5rs
1445
6205
6rs
2555
364
rs64
9330
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1420
3974
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737
rs11
3182
462
rs14
5907
211
rs17
4265
96rs
7912
2997
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rs16
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8496
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2665
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1503
7978
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1406
6622
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2433
357
rs41
4384
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7517
9742
rs76
6134
07rs
1158
7830
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1927
5324
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1411
2375
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9302
141
rs76
5478
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2017
4942
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2470
102
rs13
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861
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477
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4432
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1911
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4246
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1149
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6627
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1143
2127
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3096
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8513
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La Braña 1
rs1426654
Figure 2 | Ancestral variants around the SLC45A2 (rs16891982, above) andSLC24A5 (rs1426654, below) pigmentation genes in the Mesolithic genome.The SNPs around the two diagnostic variants (red arrows) in these two geneswere analysed. The resulting haplotype comprises neighbouring SNPs that are
also absent in modern Europeans (CEU) (n5112) but present in Yorubans(YRI) (n5113). This pattern confirms that the La Brana 1 sample is older thanthe positive-selection event in these regions. Blue, ancestral; red, derived.
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Online Content AnyadditionalMethods, ExtendedData display items and SourceData are available in the online version of the paper; references unique to thesesections appear only in the online paper.
Received 22 October; accepted 17 December 2013.
Published online 26 January 2014.
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Supplementary Information is available in the online version of the paper.
Acknowledgements The authors thank L. A. Grau Lobo (Museo de Leon) for access tothe La Brana specimen, M. Rasmussen and H. Schroeder for valid input into theexperimental work, and M. Raghavan for early access to Mal’ta genome data.Sequencing was performedat the Danish National High-Throughput DNA-SequencingCentre, University of Copenhagen. The POPRES data were obtained from dbGaP(accession number 2038). The authors are grateful for financial support from theDanish National Research Foundation, ERC Starting Grant (260372) to TM-B, and(310372) to M.G.N., FEDER and Spanish Government Grants BFU2012-38236, theSpanish Multiple Sclerosis Netowrk (REEM) of the Instituto de Salud Carlos III (RD12/0032/0011) to A.N., BFU2011-28549 to T.M.-B., BFU2012-34157 to C.L.-F., ERC(Marie Curie Actions 300554) to M.E.A., NIH NRSA postdoctoral fellowship(F32GM106656) to C.W.K.C., NIH (R01-HG007089) to J.N., NSF postdoctoralfellowship (DBI-1103639) to M.D., the Australian NHMRC to R.A.S. and a predoctoralfellowship from the Basque Government (DEUI) to I.O.
Author Contributions C.L.-F. and E.W. conceived and lead the project. M.E.P. andJ.M.V.E. provided anthropological and archaeological information. O.R. and M.E.A.performed the ancient extractions and library construction, respectively. I.O., M.E.A.,F.S.-Q., J.P.-M., S.R., O.R., M.F.-C. and T.M.-B. performed mapping, SNP calling, mtDNAassembly, contamination estimates and different genomic analyses on the ancientgenome. I.O., F.S.-Q., G.S., C.W.K.C., M.D., J.A.R., J.Q., O.R., U.M.M. and A.N. performedfunctional, ancestry and population genetic analyses. R.N. and J.N. coordinated theancestry analyses. M.G.N., R.A.S. and P.S. coordinated the immunological,pigmentation and selection analyses, respectively. I.O., M.E.A., T.M.-B., E.W. and C.L.-F.wrote themajority of themanuscriptwith critical input fromR.N.,M.G.N., J.N., R.A.S., P.S.and A.N.
Author Information Alignment data are available through the Sequence Read Archive(SRA) under accession numbers PRJNA230689 and SRP033596. Reprints andpermissions information is available at www.nature.com/reprints. The authors declareno competing financial interests. Readers are welcome to comment on the onlineversion of the paper. Correspondence and requests for materials should be addressedto E.W. ([email protected]) or C.L.-F. ([email protected]).
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Extended Data Figure 1 | Alignment and coverage statistics of the La Brana1 genome. a, Alignment summary of the La Brana 1 sequence data to hg19assembly. b, Coverage statistics per chromosome. The percentage of the
chromosome covered by at least one read is shown, as well as the mean readdepth of all positions and positions covered by at least one read. c, Percentage ofthe genome covered at different minimum read depths.
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Extended Data Figure 2 | Damage pattern of La Brana 1 sequenced reads.a, b, Frequencies of C to T (red) and G to A (blue) misincorporations at the 59
end (left) and 39 end (right) are shown for the nuclear DNA (nuDNA) (a)and mtDNA (b). c, d, Fragment length distribution of reads mapping to the
nuclear genome (c) and mtDNA genome (d). Coefficients of determination(R2) for an exponential decline are provided for the four different data sets.The exponential coefficients for the four data sets correspond to the damagefraction (l); e is the base of the natural logarithm.
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Extended Data Figure 3 | Genetic affinities of the La Brana 1 genome.a, PCA of the La Brana 1 SNP data and the 1000 Genomes Project Europeanindividuals. b, PCA of La Brana 1 versus world-wide data genotyped with theIllumina Omni 2.5M array. Continental terms make reference to each Omnipopulation grouping as follows: Africans, Yoruba and Luyha; Asians, Chinese(Beijing, Denver, South, Dai), Japanese and Vietnamese; Europeans, Iberians,
Tuscans, British, Finns and CEU; and Indian Gujarati from Texas. c, Each panelshows PC1 and PC2 based on the PCA of one of the ancient samples with themerged POPRES1FINHM sample, before Procrustes transformation. Theancient samples include the La Brana 1 sample and four Neolithic samples fromrefs 1 and 3.
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Extended Data Figure 4 | Allele-sharing analysis. Each panel shows theallele-sharing of a particular Neolithic sample from refs 1 and 3 with La Brana 1sample. The sample IDs are presented in the upper left of each panel (Ajv52,
Ajv70, Ire8, Gok4 and Otzi). In the upper right of each panel, the Pearson’scorrelation coefficient is given with the associated P value.
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Extended Data Figure 5 | Pairwise outgroup f3 statistics. a, Sardinian versus Karitiana. b, Sardinian versus Han. c, La Brana 1 versus Mal’ta. d, Sardinian versusMal’ta. e, La Brana 1 versus Karitiana. The solid line represents y 5 x.
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Extended Data Figure 6 | Analysis of heterozygosity. a, Heterozygositydistributions of La Brana 1 and modern individuals with similar coverage fromthe 1000 Genomes Project (using 1-Mb windows with 200 kb overlap). CEU,northern- and western-European ancestry. CHB, Han Chinese; FIN, Finns;
GBR, Great Britain; IBS, Iberians; JPT, Japanese; LWK, Luhya; TSI, Tuscans;YRI, Yorubans. b, Heterozygosity values in 1-Mb windows (with 200 kboverlap) across each chromosome.
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Extended Data Figure 7 | Amylase copy-number analysis. a, Sizedistribution of diploid control regions. b, AMY1 gene copy number in La Brana1. CN, copy number; DGV, Database of Genomic Variation. c, La Brana 1AMY1 gene copy number in the context of low- and high-starch dietpopulations. d, Classification of low- and high-starch diet individuals based on
AMY1 copy number. Using data from ref. 18, individuals were classified as inlow-starch (less or equal than) or high-starch (higher than) categories and thefraction of correct predictions was calculated. In addition, we calculated therandom expectation and 95% limit of low-starch-diet individuals classifiedcorrectly at each threshold value.
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Extended Data Figure 8 | Neighbouring variants for three diagnostic SNPsrelated to immunity. a, rs2745098 (PTX4 gene). b, rs11755393 (UHRF1BP1gene). c, rs10421769 (GPATCH1 gene). For PTX4, UHRF1BP1 and GPATCH1,
La Brana 1 displays the derived allele and the European-specific haplotype,indicating that the positive-selection event was already present in theMesolithic. Blue, ancestral; red, derived.
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Extended Data Figure 9 | Metagenomic analysis of the non-human reads.a, Domain attribution of the reads that did not map to hg19. b, Proportion of
different Bacteria groups. c, Proportion of different types of Proteobacteria.d, Microbial attributes of the microbes present in the La Brana 1 sample.
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