Journal of Human Genetics (2015), 1–7& 2015 The Japan Society of Human Genetics All rights reserved 1434-5161/15
www.nature.com/jhg

ORIGINAL ARTICLE

Unique characteristics of the Ainu population in
Northern Japan
Timothy A Jinam1,2,3, Hideaki Kanzawa-Kiriyama2,4, Ituro Inoue2,3, Katsushi Tokunaga5, Keiichi Omoto6
and Naruya Saitou1,2,7
Various genetic data (classic markers, mitochondrial DNAs, Y chromosomes and genome-wide single-nucleotide polymorphisms(SNPs)) have conﬁrmed the coexistence of three major human populations on the Japanese Archipelago: Ainu in Hokkaido,
Ryukyuans in the Southern Islands and Mainland Japanese. We compared genome-wide SNP data of the Ainu, Ryukyuans andMainland Japanese, and found the following results: (1) the Ainu are genetically different from Mainland Japanese living in
Tohoku, the northern part of Honshu Island; (2) using Ainu as descendants of the Jomon people and continental Asians(Han Chinese, Koreans) as descendants of Yayoi people, the proportion of Jomon genetic component in Mainland Japanese was~ 18% and ~ 28% in Ryukyuans; (3) the time since admixture for Mainland Japanese ranged from 55 to 58 generations ago,and 43 to 44 generations ago for the Ryukyuans, depending on the number of Ainu individuals with varying rates of recent
admixture with Mainland Japanese; (4) estimated haplotypes of some Ainu individuals suggested relatively long-term admixture
with Mainland Japanese; and (5) highly differentiated genomic regions between Ainu and Mainland Japanese included EDAR
and COL7A1 gene regions, which were shown to inﬂuence macroscopic phenotypes. These results clearly demonstrate theunique status of the Ainu and Ryukyuan people within East Asia.
Journal of Human Genetics advance online publication, 16 July 2015; doi:10.1038/jhg.2015.79

INTRODUCTION
The Japanese Archipelago consists of four major islands (Hokkaido,
Honshu, Shikoku and Kyushu) and many other small islands that canbe grouped into nine regions (Supplementary Figure 1). Frequent
waves of human migrations from the Eurasian continent to thearchipelago took place from at least 30 000 years ago (YBP).1 There
were various migration routes to the archipelago.2 These migrations
have shaped the human population structure in the JapaneseArchipelago, where there are currently three main populations: theAinu who mainly live in Hokkaido at the northernmost island of theArchipelago; the Ryukyuan who mainly live in the Ryukyu Islands atthe southern part; and the Mainland Japanese whose population size is
the largest and who live in all major four islands and small islands.
From an archeological perspective, the prehistory of the JapaneseArchipelago can be divided into the Paleolithic period (older than
16 000 YBP), the Jomon period (16 000–3000 YBP) and the Yayoi
period (3000–1700 YBP).1 The currently accepted model regarding theorigin of Japanese populations is the dual-structure model,3 whereby
the current Japanese population is the result of admixture between theearly migrants (Jomon people) and later migrants (Yayoi people) andthat the Ainu and the Ryukyuan are thought to retain more Jomon
components than the Mainland Japanese. Subsequent studies using1

mitochondrial DNA and several autosomal markers have been ingeneral agreement with the dual-structure model, showing theadmixed nature of Mainland Japanese4,5 and demonstrating close
afﬁnities between the Ainu and Ryukyuan populations.6,7
The Japanese Archipelago Human Population Genetics
Consortium8 produced ~ 900 000 genome-wide single-nucleotide
polymorphism (SNP) data in the Ainu and the Ryukyuans, andthrough principal component analysis (PCA), and phylogenetic tree
construction, demonstrated a clear genetic similarity between these
two groups despite their current geographical locations at the opposite
poles of the Japanese archipelago. Analysis of individual ancestry
proportions and phylogenetic analysis of the Mainland Japanese also
show that they carry both Ainu-Ryukyuan and continental Asian
genetic components. A recent study that used a model-based approach
was also in favor of the dual-structure model.9
Although previous studies generally support the dual-structure
model, some details regarding the amount of genetic contributions
from the ancestral populations were not really well deﬁned. We also
wanted to identify what kind of factors that contributed to the genetic
uniqueness that was previously observed in the Ainu.8 Therefore, theaims of this study are to perform a test for admixture and to clarify
the timing and admixture proportions in the Japanese populations.

Division of Population Genetics, National Institute of Genetics, Mishima, Japan; 2Department of Genetics, School of Life Science, Graduate University for Advanced Studies(SOKENDAI), Mishima, Japan; 3Division of Human Genetics, National Institute of Genetics, Mishima, Japan; 4Department of Anthropology, National Museum of Nature andScience, Tsukuba, Japan; 5Department of Human Genetics, Graduate School of Medicine, The University of Tokyo, Tokyo, Japan; 6Department of Anthropology, Faculty of
Science, The University of Tokyo, Tokyo, Japan and 7Department of Biological Sciences, Graduate School of Science, The University of Tokyo, Tokyo, Japan
Correspondence: Professor N Saitou, Division of Population Genetics, National Institute of Genetics, 1111 Yata, Mishima 411-0831, Japan.
E-mail: saitounr@nig.ac.jp
Received 17 November 2014; revised 9 June 2015; accepted 12 June 2015

Unique characteristics of the Ainu
TA Jinam et al
2

Table 1 Populations used for ancestry estimation tests
Population

Geographical locationnReference

Ainu

Hokkaido, Japan

36

Japanese Archipelago Population Genetics Consortium8

Ryukyuan
Mainland Japanese

Okinawa, Japan
Kanto, Japan

30
50

Japanese Archipelago Population Genetics Consortium8
Nishida et al. (2008)10

KoreanHan Chinese (CHB)

Korea
Beijing, China

50
42

Bae et al.13
The International HapMap Consortium11

European (CEU)
Yoruban (YRI)

USA
Nigeria, Africa

50
50

The International HapMap Consortium11
The International HapMap Consortium11

Malay
IndianSingapore
Singapore50
50

Teo et al.12
Teo et al.12

Han Chinese (Chinese-Sg)

Singapore50

Teo et al.12

No. of SNP loci641 314317 054906 600

570 408

Abbreviation: SNP, single-nucleotide polymorphism.

Figure 1 Principal component analysis (PCA) plot after omitting closely
related Ainu individuals.

In addition, we wish to identify highly diverged genetic loci between
the Ainu and Mainland Japanese.
MATERIALS AND METHODS
Sample data
We used 641 314 genome-wide SNP data from the Ainu and Ryukyuans
originally published by the Japanese Archipelago Human Population Genetics
Consortium8 as well as Mainland Japanese from Nishida et al.10 We merged thedata with those from three HapMap populations11 and three Singaporean
populations,12 resulting in 431 486 overlapping SNPs. We further merged thesedata with genome-wide SNP data for 50 randomly sampled Korean individuals
from Seoul who were originally used as a control group in a genome-wide
association study.13 With the inclusion of the Korean data, the number ofoverlapping SNPs was reduced to 65 256. The list of populations used in thisstudy is shown in Table 1. All of the SNP data used were already available in the
published literature.

Data analysis
Given the rather close-knit nature of the Ainu community, we investigated thepossibility that closely related individuals may be included in the data set. We
estimated measures of kinship coefﬁcients and identity-by-descent between all
pairs of Ainu individuals using REAP software,14 which can be used in
populations with admixed ancestry. It was shown previously that the Ainu haveexperienced admixture with Mainland Japanese,8 and we used individual
ancestry information at k = 2 from ADMIXTURE analysis15 (Supplementary
Figure 2) as part of input for the software.
Journal of Human Genetics

The relationship between Ainu and other populations from various
geographical locations of the Japanese archipelago was estimated using
Population Structure Prediction System for Japanese (PCAj).16 Based on theprobabilistic PCA, our genotype data were used to project individuals onto ascatterplot similar to that shown by Yamaguchi-Kabata et al.17 The Japanesesamples included in this software are part of the RIKEN Biobank collection.
To formally test whether Japanese populations are the result of
admixture between ancestral Jomon and Yayoi populations, we performed
the 3-population test (f3) using the Ainu as surrogates of Jomon ancestors andcontinental Asians (Han Chinese, Koreans) as surrogates of Yayoi ancestors.
We also performed the f4-ratio test to estimate the genetic contributions ofsource populations in the admixed populations and estimated the time since
admixture occurred using rolloff. These three tests are included in the
ADMIXTOOLS software package.18
To test whether the Ainu individuals that lie in intermediate positions
between Ainu and Mainland Japanese clusters in the PCA plot8 are recently
admixed individuals, we ﬁrst phased haplotypes in 20 Ainu individuals,
20 Mainland Japanese individuals and 8 potentially recently admixed Ainu
individuals using FastPhase19 and BEAGLE20 programs. The eight possibly
admixed Ainu individuals were phased together with the 20 Ainu and
20 Mainland Japanese separately (Supplementary Figure 3). Pairwise distances
between all phased haplotypes were calculated to generate a distance matrix thatwas used to construct a neighbor-joining tree21 to assess the afﬁnity of
haplotypes from potentially admixed Ainu individuals.
To identify genomic regions that are highly differentiated in the Ainu, we
calculated pairwise Fst22 between Ainu and Mainland Japanese after omitting
potentially recently admixed individuals from each population. We focused on
the top 1% of highly differentiated SNPs and performed gene annotation searchusing GOrilla gene ontology tool23 to ﬁnd out whether these SNPs have any
signiﬁcant biological functions.

RESULTS
We identiﬁed ﬁve parent-offspring and two sibling pairs in the Ainu
based on the values of kinship coefﬁcient and probability of identityby-descent = 0 (Supplementary Figure 4). We therefore omitted one
individual from each of the parent–offspring pairs, and used theremaining 31 individuals for PCA. The result of the new PCA using
approximately 65 k SNPs is shown in Figure 1. The ﬁrst principal
component (PC1) separates the Ainu from the rest of East Asian
populations, and the population closest to the Ainu are Ryukyuans,consistent with the previous observations. The Ainu individuals are
spread out in the same ‘comet-like’ pattern as before,8 but no outlier
Ainu individuals were distinguished from PC2. When the Korean data
set was omitted, the resulting PCA using about 430k SNPs also showeda similar pattern (Supplementary Figure 5). We therefore surmise thatthe ﬁve outlier Ainu individuals seen in the previous PCA plot8
represent an artifact due the inclusion of very closely related
individuals within the Ainu population.

Unique characteristics of the Ainu
TA Jinam et al3

The evolutionary factors that created variations explained by PC2
are difﬁcult to conjecture. If we disregard Ryukyuan individuals, thenthe PC2 axis from top to bottom seems to reﬂect a south to northgeographical cline in populations: Singapore Han Chinese, Beijing Han
Chinese (CHB), Koreans and Mainland Japanese of the JapaneseArchipelago. However, Ryukyuan individuals are located on the top

Figure 2 Scatterplot output from PCAj software. The three populations
from Japanese Archipelago Human Population Genetics Consortium8 areshown in colored dots. Other Japanese and Chinese individuals are shown asgray dots.

part on the PC2 axis, above the Mainland Japanese of the JapaneseArchipelago. A possible explanation would be some unknown
populations were involved in the formation of Ryukyuans, as
previously suggested.24
We identiﬁed eight Ainu individuals who might be recently
admixed with Mainland Japanese based on their intermediate
positions between Ainu and Mainland Japanese clusters in the PCA
plot (Supplementary Figure 3). If these individuals were the result ofvery recent admixture events, then one of the pair of chromosomes
should be from an Ainu parent and the other from a Mainland
Japanese parent. The neighbor joining tree of chromosome 22
haplotypes that were phased using Fastphase and BEAGLE for eachof the eight admixed Ainu individuals is shown in Supplementary
Figure 6. The haplotype afﬁnities for these potentially recently
admixed Ainu did not show a consistent pattern, with three
individuals having haplotype afﬁnities with the Ainu and anotherfour individuals with afﬁnities to Mainland Japanese haplotypes.
However, in one individual (labeled 2120001B03), one of the
haplotypes clustered with other Ainu haplotypes while the otherclustered with Mainland Japanese haplotypes, indicating that thisperson might be a result of recent admixture.
The 31 Ainu individuals together with 35 Ryukyuan and 50
randomly chosen Mainland Japanese from the Kanto region werecompared with other individuals from various geographical locationsin the Japanese archipelago using probabilistic PCA.16 Our samples
were overlaid on a scatterplot that showed major clustering between
Mainland Japanese, CHB and Ryukyuans. Our Ryukyuan (green dots)
and Mainland Japanese samples from the Kanto region (red dots) fallwithin well-deﬁned clusters reported by Yamaguchi-Kabata et al.17 as
seen in Figure 2. Interestingly, the Ainu individuals (blue dots) form a
gradient alongside the cluster of Ryukyuan individuals on the verticalaxis. There are several individuals from the RIKEN data set (identiﬁed
as gray dots) that cluster with our Ainu samples. Although their
speciﬁc ethnicity was not directly mentioned, they are most likely Ainu
individuals based on their geographical origin, which is Hokkaido
where most Ainu people currently reside. This identiﬁcation of Ainu
people in the RIKEN data was not reported by Yamaguchi-Kabata
et al.17 nor Kumasaka et al.16
We tested the dual-structure model for the origin of modern
Japanese using the 3-population test (f3 test). We used two data sets

Table 2 Results of 3-population admixture test
Source population 1
Using ~ 400 k SNP data set
Ainu

Target population

Source population 2

f3

Standard error

Z-scoreMainland Japanese

CHB− 0.0070

0.000242− 28.8Ainu
Ainu

Mainland Japanese
RyukyuanChinese-Sg
CHB− 0.0068− 0.0089

0.000258
0.000359− 26.5− 24.9Ainu
Ainu

Ryukyuan
RyukyuanChinese-Sg
Mainland Japanese− 0.0091− 0.0046

0.000376
0.000278− 24.1− 16.5

Ainu
Ainu

Mainland Japanese
Mainland Japanese

KoreanCHB− 0.0071− 0.0066

0.000248
0.000271− 28.6− 24.2

Ainu
Ainu

Mainland Japanese
RyukyuanChinese-Sg
Korean− 0.0064− 0.0089

0.000283
0.000363− 22.8− 24.7

Using ~ 60 k SNP with the inclusion of Korean data

Ainu

Ryukyuan

CHB− 0.0084

0.000367− 22.9Ainu
Ainu

Ryukyuan
RyukyuanChinese-Sg
Mainland Japanese− 0.0086− 0.0043

0.000377
0.000296− 22.7− 14.7Abbreviations: CHB, Beijing Han Chinese; SNP, single-nucleotide polymorphism.

Journal of Human Genetics

Unique characteristics of the Ainu
TA Jinam et al4

for this test: a high density SNP data set without Korean data (~430 k
SNPs) and a low density data set with Korean data (~65 k SNPs).
Negative values for the f3 test imply that the target population is theresult of admixture between two source populations. The Z-score forthe test can be taken as a measure of statistical signiﬁcance. The
combination using continental Asians (Han Chinese, Koreans) andAinu as source populations and the Mainland Japanese and Ryukyuan
as target populations showed the most signiﬁcant results (Table 2).
Using the high density SNP data set, the (Mainland Japanese; CHB,
Ainu) combination gave the most signiﬁcant result (f3 = − 7.0 × 10 − 3;Z-score = − 28.8), whereas in the low density SNP data set, thecombination (Mainland Japanese; Korean, Ainu) had the mostsigniﬁcant result (f3 = − 7.1 × 10 − 3; Z-score = − 28.6). Tests using
Ryukyuans as target populations also showed similar patterns, but withlower Z-scores compared with Mainland Japanese. These results showedthat the Mainland Japanese were the result of admixture between theancestors of Han Chinese/Koreans and Ainu, who represent the
Yayoi and Jomon peoples, respectively. This adds further support tothe dual-structure model3 for the origin of Mainland Japanese.
To estimate the proportion of genetic contributions from theancestral populations in the Japanese, we conducted the f4-ratio
estimation test. Populations used in this test are assumed to be related
to each other according to the tree shown in Figure 3. This test differsfrom the f3 test in the inclusion of an outgroup, and anotherpopulation that is close to one of the source populations (continental

Asians). Using various combinations of populations, the results withthe most signiﬁcant Z-scores are shown in Table 3. The proportion of
the Jomon ancestry (ancestors of Ainu) in Mainland Japanese wasestimated to be 17.8% (Z-score 72.3) when using the CHB as the othersource population. When using the smaller data set with Koreansas the source population, the proportion of the Jomon ancestryin Mainland Japanese was higher at 17.9% (Z-score 64.9). The
proportion of Jomon components in the Ryukyuans was from
28.4% (Z-score 43.8) when using high density SNP data and 27.8%(Z-score 40.0) when using the lower density SNP data. The higher
proportion of Jomon component in the Ryukyuans compared with theMainland Japanese was consistent with the individual ancestry
estimates using ADMIXTURE (Supplementary Figure 2).
We further estimated the time since the admixture event between
the Yayoi and Jomon ancestors that resulted in the Japanese
populations using the rolloff program.18 Due to the presence ofadmixed individuals within Ainu, we decided to create three subsets of
Ainu individuals with different levels of admixture (Supplementary
Figure 7) to gauge the performance of the rolloff program. Because
this test is based on the decay of linkage disequilibrium over time, weused the higher density SNP data set (~430 k SNP). Thus, we took
CHB and the three Ainu subsets as ancestral populations, and theMainland Japanese and Ryukyuan as admixed populations. The resultsin Table 4 show that the Ainu data set with the least number ofadmixed individuals (Ainu-15) yielded a much older time since

Table 4 Estimation of time since admixture for Mainland Japaneseand Ryukyuans, using Han Chinese (CHB) and Ainu as source
populations
Time since admixture in
generations (years)a
Ainu ancestry (admixture
Ainu dataset

Figure 3 Schematic diagram showing the relationship between populations
used for the estimation of Jomon (1 − α) and Yayoi (α) proportions in the
Mainland Japanese.

results at k = 2) (%)

Mainland Japanese

Ryukyuan

Ainu-15
Ainu-2098.2
92.9

58 (1450)
56 (1400)

44 (1100)
43 (1075)

Ainu-2882.5

55 (1375)

43 (1075)

Three Ainu data sets with different proportions of admixture were used.
aA generation time of 25 years was used.

Table 3 Proportion of Jomon ancestry (1 − α) estimated from the f4-ratio testco(1 − α)Standard error

Z-scoreMainland Japanese
Mainland Japanese

Ainu
Ainu

IndianIndian0.17800.13630.01140.012872.3
67.3

Mainland Japanese
Mainland Japanese

Ryukyuan
Ryukyuan

Ainu
Ainu

IndianIndian0.28450.27170.01640.0167

43.8
43.7

CHB
Chinese-Sg

KoreanKoreanMainland Japanese
Mainland Japanese

Ainu
Ainu

IndianIndian0.17940.17440.01260.0129

64.9
63.8

CHB
Chinese-Sg

Chinese-Sg
CHB

Mainland Japanese
Mainland Japanese

Ainu
Ainu

IndianIndian0.13920.19600.01470.013858.7
58.2

KoreanMainland Japanese

Ryukyuan

Ainu

Indian0.27880.018040.0

Chinese-Sg
CHB

Mainland Japanese
Mainland Japanese

Ryukyuan
Ryukyuan

Ainu
Ainu

IndianIndian0.27440.29450.01900.0190

38.2
37.2

a

b

x

Chinese-Sg
CHB

CHB
Chinese-Sg

CHB
Chinese-Sg

Using ~ 400 k SNP data set

Using ~ 60 k SNP with the inclusion of Korean data

Abbreviations: CHB, Beijing Han Chinese; SNP, single-nucleotide polymorphism. Populations labels (a, b, x, c, o) are ordered as in Figure 3.

Journal of Human Genetics

Unique characteristics of the Ainu
TA Jinam et al5

admixture (58 generations ago or 1450 years ago, assuming ageneration time of 25 years) when using Mainland Japanese as the
admixed population. With the inclusion of more admixed individuals
in the Ainu data set, the time since admixture became more recent(55 generations ago or 1375 years ago). The estimates using
Ryukyuans as the admixed population ranged from 44 to 43
generations ago (1100–1075 years ago), which are relatively more
recent compared with those for the Mainland Japanese.
We also identiﬁed SNP loci that are differentiated between the Ainu
and Mainland Japanese by using pairwise Fst values. The pairwise Fst
values ranged from 0 to 0.8903, with a mean of 0.0407. The majorityof the SNPs (approximately 400 000) have Fst values of less than 0.02.
We picked 6413 SNPs that were within the top 1% and had Fst values
higher than 0.36. The Fst values and annotations for these top 1% SNP
are listed in Supplementary Table 1. Within those top 1% of SNP,
some of them were found in genes reported to be associated with facial
structure in Europeans25 and hair and tooth morphology in East
Asians.26,27 The distribution of Fst values for SNPs in those genes areshown in Figure 4. Two out of ﬁve genes for facial morphology (PAX3and COL17A1) contain highly differentiated SNPs, as with the hair/
tooth morphology gene (EDAR). The results of gene annotation
analysis on those top 1% SNPs showed enrichment for biologicalprocesses and cellular components involving collagen (Supplementary
Figure 8).

DISCUSSION
We identiﬁed several very closely related pairs of individuals within the
Ainu, which was not reported previously.8 The inclusion of closely
related individuals may produce artifactual results for individual-based
clustering tests such as PCA or STRUCTURE. Although the PCA plot
after omission of closely related Ainu individuals is slightly different
from the previous result,8 the overall pattern remains unchanged. The
gradient of Ainu individuals along PC1 can be explained partially byrecent admixture between Ainu and Mainland Japanese parents andalso intermarriage between individuals with different proportions of
Ainu and Mainland Japanese ancestry.
We also had the opportunity to compare our Ainu data with thosefrom other Japanese living in different locations in the JapaneseArchipelago that was represented in the RIKEN Biobank data.16
The Ryukyuans and Mainland Japanese from Kanto region(Supplementary Figure 1) in our data overlap with the RIKEN data.
Ainu individuals still form the gradient, but are positioned adjacent tothe cluster of Ryukyuan populations (Figure 2). The three Ainu
individuals appearing in the Mainland Japanese cluster were also
observed with the RIKEN data. Although there was no major
discrepancy between our data and the RIKEN data, the position of
the Ainu in Figure 2 is slightly different from Figure 1. This may
reﬂect biases in sample sizes between the two data sets. In the RIKEN
data, the majority of the individuals (approximately 1000) are from

Figure 4 Distribution of differentiated SNP in the Ainu for the ﬁve genes associated with facial morphology in Europeans (Liu et al.25) and for hair and toothmorphology (EDAR). The top 1% highly differentiated SNPs have Fst values above 0.36, represented by the red horizontal lines.
Journal of Human Genetics

Unique characteristics of the Ainu
TA Jinam et al6

the Mainland Japanese cluster, with only a few individuals placed
within our Ainu cluster.
A clear difference between the northern Tohoku region Mainland
Japanese and Ainu is another important ﬁnding. Yamaguchi-Kabata
et al.17 showed that people living in the Tohoku region are distributed
slightly downward in the Mainland Japanese cluster, which overlaps
with Kanto region Mainland Japanese (red color) shown in Figure 2.
Historically, people in northern Tohoku area were called ‘Emishi’,while the Ainu people were called ‘Ezo’, using the same Chinesecharacters. It is still not clear whether the Emishi and Ezo people were
genetically similar. Some cultural components such as locale names andpotteries are shared between the Emishi in Tohoku region and the Ezo
in Hokkaido who eventually evolved into the Ainu people.28 However,the PCA shown in Figure 2 suggests that some unknown factor(s) may
exist in the formation of Emishi people in the Tohoku region.
Japanese Archipelago Human Population Genetics Consortium8
presented a ﬁgure that represents the possible evolutionary historyof populations in the Japanese Archipelago. Here we did a formal test
using the 3-population, f4 ratio estimation and rolloff tests that has
been previously used in other populations.29 In particular, we wanted
to test the plausibility of the dual-structure model for the origin of
current Japanese using the Ainu and continental Asians as ancestral
populations. The results of the 3-population test show that theMainland Japanese are indeed a result of admixture between theancestors of the Ainu (Jomon people) and continental Asians (Yayoi
People), with an estimated 18% contribution from the Jomon
ancestry. Results using the Ryukyuans also indicate that they are
admixed, although the Z-scores were less signiﬁcant when using
Mainland Japanese (Tables 2 and 3). The estimated Jomon component
was higher in the Ryukyuans, at approximately 28%. This was
expected, given the low but signiﬁcant afﬁnity between the Ainu
and the Ryukyuans. Our Jomon ancestry estimates were lower thanwhat was reported previously, which ranged from 30 to 40% in the
Mainland Japanese and 50–60% in the Ryukyuans.30,31 In this study,
we used high density autosomal SNP data in the Ainu, whereas
previous studies used only Y-chromosome polymorphisms,30 or using
non-Ainu as the ancestral population.31 These differences in addition
to other methodological parameters may lead to the differences in theestimates.
The time since admixture estimated using rolloff varied from 55 to
58 generations ago, depending on the number of admixed Ainu
included in the data set. If we had more Ainu individuals who are lessadmixed with Mainland Japanese, then the estimate could be evenolder. Our estimated time since admixture of 1450 years ago may betreated as a lower bound, since rolloff assumes only one admixture
event and estimates the most recent event. This time estimate
coincides with the period, during 5th–8th century, in which theJapanese government expanded their territory to the northern part
(Tohoku) of the Honshu Island.28 Interestingly, the time since theadmixture event that led to the formation of Ryukyuans was more
recent compared with the Mainland Japanese. The estimated admixture time of approximately 1100 years ago corresponds to the Gusuku
period of Okinawan archeological history.32 A plausible explanation
may involve the following scenario: the Jomon people who alreadysettled in Ryukyu Islands experienced admixture with migrants fromthe continent who themselves may have already admixed with the
Jomon people in the Japanese Mainland.
We also report several SNP loci that are highly differentiatedbetween the Ainu and the Mainland Japanese. These include two genes
associated with facial structure in Europeans.25 SNPs within the EDAR
gene have been reported to be highly differentiated between Mainland
Journal of Human Genetics

Japanese and Ryukyuans.17 Similar patterns were observed whencomparing between Ainu and Mainland Japanese. However, thenon-synonymous SNP rs3827760 reported by Yamaguchi-Kabata
et al.17 was not covered on the Affymetrix 6.0 genechip. They also
reported a SNP within the ABCC11 gene that was responsible forearwax type was highly differentiated between the Ryukyuan and theMainland Japanese. However, possibly due to poor coverage of the
ABCC11 gene in the Affymetrix genechip, all SNPs in that gene were
found to be monomorphic between the Ainu and the Mainland
Japanese. It is known that the Ainu have very different physicalcharacteristics compared with Mainland Japanese, and the ﬁnding ofdifferentiated loci associated with observable phenotypes agrees withthe works done by physical anthropologists over the years.33
To conclude, we supplemented our initial analysis on the Ainu
population, and demonstrated that their ancestors contributed
signiﬁcantly to the genetic makeup of the current populations on
the Japanese Archipelago. If we consider a very unique genetic status
of Jomon people, then it is understandable that Ainu people, who
inherited the highest proportion of Jomon DNA, are quite uniqueamong all extant East Eurasians.
CONFLICT OF INTEREST
The authors declare no conﬂict of interest.
ACKNOWLEDGEMENTS
We thank Japanese Archipelago Human Population Genetics Consortium andAsian DNA Repository Consortium for permission of access to SNP data. This
study was supported by a SOKENDAI Grant on modern human dispersal.
Finally, we appreciate people on the Japanese Archipelago (Ainu, Ryukyuan and
Mainland Japanese) who kindly donated their DNA to this and past
anthropological studies.

1 Imamura, K. Prehistoric Japan: New Perspectives on Insular East Asia (University of
Hawaii Press, Honolulu, HI, USA, 1996).
2 Saitou, N. in Simulations, Genetics and Human Prehistory (eds Matsumura S., Forster P.
& Renfrew C.) 89–92 (McDonald Institute Monographs, Cambridge, UK, 2008).
3 Hanihara, K. Dual structure model for the population history of the Japanese. Japan
Rev. 2, 1–33 (1991).
4 Tanaka, M., Cabrera, V. M., González, A. M., Larruga, J. M., Takeyasu, T., Fuku, N.
et al. Mitochondrial genome variation in eastern Asia and the peopling of Japan.
Genome Res. 14, 1832–1850 (2004).
5 Nakaoka, H., Mitsunaga, S., Hosomichi, K., Shyh-yuh, L., Sawamoto, T., Fujiwara, T.
et al. Detection of ancestry informative HLA alleles conﬁrms the admixed origins ofJapanese population. PLoS ONE 8, e60793 (2013).
6 Omoto, K. & Saitou, N. Genetic origins of the Japanese: a partial support for the dual
structure hypothesis. Am J Phys Anthropol. 102, 437–446 (1997).
7 Bannai, M., Ohashi, J., Harihara, S., Takahashi, Y., Juji, T., Omoto, K. et al. Analysis of
HLA genes and haplotypes in Ainu (from Hokkaido, northern Japan) supports thepremise that they descent from Upper Paleolithic populations of East Asia. Tissue
Antigens 55, 128–139 (2000).
8 Japanese Archipelago Human Population Genetics Consortium. [Jinam, T., Nishida, N.,
Hirai, M., Kawamura, S., Umetsu, K. et al.] The history of human populations in theJapanese Archipelago inferred from genome-wide SNP data with a special reference to
the Ainu and the Ryukyuan populations. J. Hum. Genet. 57, 787–795 (2012).
9 Nakagome, S., Sato, T., Ishida, H., Hanihara, T., Yamaguchi, T., Kimura, R. et al.
Model-based veriﬁcation of hypotheses on the origin of modern Japanese revisited by
Bayesian inference based on genome-wide SNP data. Mol. Biol. Evol. 32,
1533–1543 (2015)
10 Nishida, N., Koike, A., Tajima, A., Ogasawara, Y., Ishibashi, Y., Uehara, Y., et al.
Evaluating the performance of Affymetrix SNP Array 6.0 platform with 400 Japaneseindividuals. BMC Genomics 9, 431–431 (2008).
11 The International HapMap Consortium. A haplotype map of the human genome. Nature
437, 1299–1320 (2005).
12 Teo, Y. Y., Sim, X., Ong, R. T., Tan, A. K., Chen, J., Tantoso, E. et al. Singapore
Genome Variation Project: a haplotype map of three Southeast Asian populations.
Genome Res. 19, 2154–2162 (2009).
13 Bae, J. S., Cheong, H. S., Kim, J. H., Park, B. L., Kim, J. H., Park, T. J. et al. The
genetic effect of copy number variations on the risk of type 2 diabetes in a Korean
population. PLoS ONE 6, e19091 (2011).

Unique characteristics of the Ainu
TA Jinam et al7
14 Thornton, T., Tang, H., Hoffman, T. J., Ochs-Balcom, H. M., Baan, B. J. & Risch, N.
Estimating kinship in admixed populations. Am. J. Hum. Genet. 91, 122–138 (2012).
15 Alexander, D. H., Novembre, J. & Lange, K. Fast model-based estimation of ancestry inunrelated individuals. Genome Res. 19, 1655–1664 (2009).
16 Kumasaka, N., Yamaguchi-Kabata, Y., Takahashi, A., Kubo, M., Nakamura, Y. &Kamatani, N. Establishment of a standardized system to perform population structure
analyses with limited sample size or with different sets of SNP genotypes. J. Hum.
Genet. 55, 525–533 (2010).
17 Yamaguchi-Kabata, Y., Nakazono, K., Takahashi, A., Saito, S., Hosono, N., Kubo, M.
et al. Population structure of Japanese based on SNP genotypes from 7,001 individuals
in comparison to other ethnic groups: Effects on population-based association studies.
Am. J. Hum. Genet. 83, 445–456 (2008).
18 Patterson, N., Moorjani, P., Luo, Y., Mallick, S., Rohland, N., Zhan, Y. et al. Ancient
admixture in human history. Genetics 192, 1065–1093 (2012).
19 Scheet, P. & Stephens, M. A fast and ﬂexible statistical model for large-scale population
genotype data: applications to inferring missing genotypes and haplotypic phase. Am. J.
Hum. Genet. 78, 629–644 (2006).
20 Browning, S. R. & Browning, B. L. Rapid and accurate haplotype phasing and missing
data inference for whole genome association studies by use of localized haplotype
clustering. Am. J. Hum. Genet. 81, 1084–1097 (2007).
21 Saitou, N. & Nei, M. The neighbor-joining method: a new method for reconstructing
phylogenetic trees. Mol. Biol. Evol. 4, 406–425 (1987).
22 Weir, B. S. & Cockerham, C. C. Estimating F-statistics for the analysis of population
structure. Evolution 38, 1358–1370 (1984).
23 Eden, E., Navon, R., Steinfeld, I., Lipson, D. & Yakhini, Z. GOrilla: a tool for discovery
and visualization of enriched GO terms in ranked gene lists. BMC Bioinformatics 10,
48 (2009).

24 Sato, T., Nakagome, S., Watanabe, C., Yamaguchi, K., Kawaguchi, A., Koganebuchi, K.
et al. Genome-Wide SNP analysis reveals population structure and demographic historyof the Ryukyu Islanders in the Southern part of the Japanese Archipelago. Mol. Biol.
Evol. 31, 2929–2940 (2014).
25 Liu, F., van der Lijn, F., Schurmann, C., Zhu, G., Chakravarty, M. M., Hysi, P. G. et al.
A genome-wide association study identiﬁes ﬁve loci inﬂuencing facial morphology inEuropeans. PLoS Genet. 8, e1002932 (2012).
26 Fujimoto, A., Kimura, R., Ohashi, J., Omi, K., Yuliwulandari, R., Batubara, L. et al.
A scan for genetic determinants of human hair morphology: EDAR is associated with
Asian hair thickness. Hum. Mol. Genet 17, 835–843 (2008).
27 Kimura, R., Yamaguchi, T., Takeda, M., Kondo, O., Toma, T., Haneji, K. et al.
A common variation in EDAR is a genetic determinant of shovel-shaped incisors. Am. J.
Hum. Genet. 85, 528–535 (2009).
28 Kudo, M. Ancient History of Emishi, (Heibon-sha, Tokyo, Japan, 2001) [in Japanese].
29 Moorjani, P., Thangaraj, K., Patterson, N., Lipson, M., Loh, P. R., Govindaraj, P. et al.
Genetic evidence for recent population mixture in India. Am. J. Hum. Genet. 93,
422–438 (2013).
30 Rasteiro, R. & Chikhi, L. Revisiting the peopling of Japan: an admixture perspective.
J. Hum. Genet. 54, 349–354 (2009).
31 He, Y., Wang, W. R. & Xu, S.The HUGO Pan-Asian SNP Consortium. Paleolithic
contingent in modern Japanese : estimation and inference using genome-wide data.
Sci. Rep. 2, 1–7 (2012).
32 Tomiyama, K. World of Ryukyu and Okinawa History, Yoshikawa Kobunkan, Tokyo(2003) [in Japanese].
33 Ossenberg, N. S., Dodo, Y., Maeda, T. & Kawakubo, Y. Ethnogenesis and craniofacialchange in Japan from the perspective of nonmetric traits. Anthropol. Sci. 114,
99–115 (2006).

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