The Date of Interbreeding between Neandertals and
Sriram Sankararaman1,2*, Nick Patterson2, Heng Li2, Svante Pa ¨a ¨bo3*, David Reich1,2*
1Department of Genetics, Harvard Medical School, Boston, Massachusetts, United States of America, 2Broad Institute of MIT and Harvard, Cambridge, Massachusetts,
United States of America, 3Department of Evolutionary Genetics, Max Planck Institute for Evolutionary Anthropology, Leipzig, Germany
Comparisons of DNA sequences between Neandertals and present-day humans have shown that Neandertals share more
genetic variants with non-Africans than with Africans. This could be due to interbreeding between Neandertals and modern
humans when the two groups met subsequent to the emergence of modern humans outside Africa. However, it could also
be due to population structure that antedates the origin of Neandertal ancestors in Africa. We measure the extent of linkage
disequilibrium (LD) in the genomes of present-day Europeans and find that the last gene flow from Neandertals (or their
relatives) into Europeans likely occurred 37,000–86,000 years before the present (BP), and most likely 47,000–65,000 years
ago. This supports the recent interbreeding hypothesis and suggests that interbreeding may have occurred when modern
humans carrying Upper Paleolithic technologies encountered Neandertals as they expanded out of Africa.
Citation: Sankararaman S, Patterson N, Li H, Pa ¨a ¨bo S, Reich D (2012) The Date of Interbreeding between Neandertals and Modern Humans. PLoS Genet 8(10):
Editor: Joshua M. Akey, University of Washington, United States of America
Received December 15, 2011; Accepted July 27, 2012; Published October 4, 2012
Copyright: ? 2012 Sankararaman et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits
unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
Funding: This work was supported by the Presidential Innovation Fund of the Max Planck Society, the Krekeler Foundation, and the National Science Foundation
(HOMINID grant 1032255). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.
Competing Interests: The authors have declared that no competing interests exist.
* E-mail: firstname.lastname@example.org (SS); email@example.com (SP); firstname.lastname@example.org (DR)
A much-debated question in human evolution is the relationship
between modern humans and Neandertals. Modern humans
appear in the African fossil record about 200,000 years ago.
Neandertals appear in the European fossil record about 230,000
years ago  and disappear about 30,000 year ago. They lived in
Europe and western Asia with a range that extended as far east as
Siberia  and as far south as the middle East. The overlap of
Neandertals and modern humans in space and time suggests the
possibility of interbreeding. Evidence, both for  and against
interbreeding , have been put forth based on the analysis of
modern human DNA. Although mitochondrial DNA from
multiple Neandertals has shown that Neandertals fall outside the
range of modern human variation [5,6,7,8,9,10], low-levels of
gene flow cannot be excluded [10,11,12].
Analysis of the draft sequence of the Neandertal genome
revealed that the Neandertal genome shares more alleles with non-
African than with sub-Saharan African genomes . One
hypothesis that could explain this observation is a history of gene
flow from Neandertals into modern humans, presumably when
they encountered each other in Europe and the Middle East 
(Figure 1). An alternative hypothesis is that the findings are
explained by ancient population structure in Africa [13,14,15,16],
whereby the population ancestral to Neandertal and modern
human ancestors was subdivided. If this substructure persisted
until modern humans carrying Upper Paleolithic technologies
expanded out of Africa so that the modern human population that
migrated was genetically closer to Neandertals, people outside
Africa today would share more genetic variants with Neandertals
that people in sub-Saharan Africa [13,14,15] (Figure 1). Ancient
substructure in Africa is a plausible alternative to the hypothesis of
recent gene flow. Today, sub-Saharan Africans harbor deep
lineages that are consistent with a highly-structured ancestral
population [17,18,19,20,21,22,23,24,25,26,27]. Evidence for an-
cient structure in Africa has also been offered based on the
substantial diversity in neurocranial geometry amongst early
modern humans . Thus, it is important to test formally
whether substructure could explain the genetic evidence for
Neandertals being more closely related to non-Africans than to
A direct way to distinguish the hypothesis of recent gene flow
from the hypothesis of ancient substructure is to infer the date for
when the ancestors of Neandertals and a modern non-African
population last exchanged genes. In the recent gene flow scenario,
the date is not expected to be much older than 100,000 years ago,
corresponding to the time of the earliest documented modern
humans outside of Africa . In the ancient substructure
scenario, the date of last common ancestry is expected to be at
least 230,000 years ago, since Neandertals must have separated
from modern humans by that time based on the Neandertal fossil
record of Europe .
In present-day human populations, the extent of LD between
two single nucleotide polymorphisms (SNPs) shared with Nean-
dertals can be the result of two phenomena. First, there is ‘‘non-
admixture LD’’  whose extent reflects stretches of DNA
inherited from the ancestral population of Neandertals and
modern humans as well as LD that has arisen due to bottlenecks
and genetic drift in modern humans since they separated from
Neandertals. Second, if gene flow from Neandertals into modern
humans occurred, there is ‘‘admixture LD’’ , which will reflect
stretches of genetic material inherited by modern humans through
PLOS Genetics | www.plosgenetics.org1 October 2012 | Volume 8 | Issue 10 | e1002947
interbreeding with Neandertals. The extent of LD between single
nucleotide polymorphisms (SNPs) shared with Neandertals will
thus reflect, at least in part, the time since Neandertals or their
ancestors and modern humans or their ancestors last exchanged
genes with each other.
The strategy of using LD to estimate dates of gene flow events
[31,32,33,34,35]. Our methodology is conceptually similar to the
methodology developed by Moorjani et al., but is dealing with a
more challenging technical problem since the methodology
developed by Moorjani et al. is adapted for relatively recent
admixtures. In recently admixed populations that have not
experienced recent bottlenecks, admixture LD extends over size
scales at which non-admixture LD makes a negligible contribu-
tion. Thus, one can infer the time of gene flow based on inter-
marker spacings that are larger than the scale of non-admixture
LD. For older admixtures however (such as may have occurred in
the case of Neandertals), non-admixture LD occurs almost at the
same size scale as admixture LD. To account for this, we study
pairs of markers that are very close to each other, but ascertain
them in a way that greatly minimizes the signals of non-admixture
LD while enhancing the signals of admixture LD. Thus, unlike in
the case of recent admixtures, non-admixture LD could bias an
admixture date obtained using our methods; however, we show
using simulations of a very wide set of demographic scenarios that
our marker ascertainment procedure makes the bias so small that
our inferences are qualitatively unaffected.
Our methodology is based on the idea that if two alleles, a
genetic distance x (expected number of crossover recombination
events per meiosis) apart, arose on the Neandertal lineage and
introgressed into modern humans at time tGF, the probability that
these alleles have not been broken up by recombination since gene
explored byseveral groups
Figure 1. Linkage disequilibrium patterns expected due to recent gene flow and ancient structure. (A) In the case of recent gene flow
from Neandertals (NEA) into the ancestors of non-Africans (CEU) but not into the ancestors of Africans (YRI), we expect long range LD at sites where
Neandertal has the derived allele, and this expectation of admixture generated LD is verified by computer simulation as shown in the right of the
panel along with a fitted exponential decay curve. (B) In the case of ancient structure, we expect short range LD, reflecting the time since Neandertals
and non-Africans derived from a shared ancestral population, and this expectation is also verified by simulation.
One of the key discoveries from the analysis of the
Neandertal genome is that Neandertals share more genetic
variants with non-Africans than with Africans. This obser-
vation is consistent with two hypotheses: interbreeding
between Neandertals and modern humans after modern
humans emerged out of Africa or population structure in
the ancestors of Neandertals and modern humans. These
hypotheses make different predictions about the date of
last gene exchange between the ancestors of Neandertals
and modern non-Africans. We estimate this date by
measuring the extent of linkage disequilibrium (LD) in
the genomes of present-day Europeans and find that the
last gene flow from Neandertals into Europeans likely
occurred 37,000–86,000 years before the present (BP), and
most likely 47,000–65,000 years ago. This supports the
recent interbreeding hypothesis and suggests that inter-
breeding occurred when modern humans carrying Upper
Paleolithic technologies encountered Neandertals as they
expanded out of Africa.
Dating Neandertal Gene Flow into Modern Humans
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Dating Neandertal Gene Flow into Modern Humans
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