Hindawi Publishing Corporation
Journal of Biomedicine and Biotechnology
Volume 2012, Article ID 789024, 10 pages
doi:10.1155/2012/789024

Research Article
Copy-Number Variations Observed in a Japanese Population by
BAC Array CGH: Summary of Relatively Rare CNVs
Yasunari Satoh,1 Keiko Sasaki,1 Yuko Shimoichi,1 Keiko Sugita,2Hiroaki Katayama,2 and Norio Takahashi1
1 Department of Genetics, Radiation Eﬀects Research Foundation, 5-2 Hijiyama Park, Minami-ku, Hiroshima 732-0815, Japan
2 Department of Information Technology, Radiation Eﬀects Research Foundation, 5-2 Hijiyama Park, Minami-ku,

Hiroshima 732-0815, Japan
Correspondence should be addressed to Norio Takahashi, takahash@rerf.or.jp
Received 6 July 2011; Revised 21 September 2011; Accepted 7 October 2011
Academic Editor: Xin-yuan Guan
Copyright © 2012 Yasunari Satoh et al. This is an open access article distributed under the Creative Commons Attribution License,
which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.
Copy-number variations (CNVs) may contribute to genetic variation in humans. Reports regarding existence and characteristics of
CNVs in a large apparently healthy Japanese cohort are quite limited. We report the data from a screening of 213 unrelated Japaneseindividuals using comparative genomic hybridization based on a bacterial artificial chromosome microarray (BAC aCGH). In aprevious paper, we summarized the data by focusing on highly polymorphic CNVs (in ≥5.0 % of the individuals). However, rarevariations have recently received attention from scientists who espouse a hypothesis called “common disease and rare variants.”
Here, we report CNVs identified in fewer than 10 individuals in our study population. We found a total of 126 CNVs at 52 diﬀerent
BAC regions in the genome. The CNVs observed at 27 of the 52 BAC-regions were found in only one unrelated individual. The
majority of CNVs found in this study were not identified in the Japanese who were examined in the other studies. Family studies
were conducted, and the results demonstrated that the CNVs were inherited from one parent in the families.

1. Introduction
Genomes vary from one another in multifarious ways, andthe totality of this genetic variation underpins the heritabilityof human traits. Various recent studies show the landscapeof genetic variation and allow estimation of the relative contributions of sequence (base substitutions) and structuralvariation (indels, insertions, or deletions; copy-number variations [CNVs], and inversions). Among DNA sequence variations in the human genome, CNVs directly contribute togene expression changes through gene dosage eﬀects; the distribution of CNVs is now considered to be wider than previously thought [1–6]. Genome resequencing studies haveshown that CNVs, involving the gain or loss of several hundreds of bases to several hundred kilobases (kb) of the genome, can be an important source of genetic variation among
human populations of diﬀerent ethnic groups as well as
among individuals. Following the development of methodologies and the introduction of new research platforms [7–12], information regarding the nature and pattern of CNVs

from representative populations has been accumulated. The
functional impact of CNVs has been demonstrated across thefull range of biology [13], from cellular phenotypes, such as
gene expression [14], to all classes of human disease with anunderlying genetic basis: sporadic, Mendelian, complex, andinfectious (reviewed in [15]).
Clinical geneticists need to discriminate pathogenic frombenign CNVs in their patients and have made extensive useof data from CNV surveys of apparently healthy individuals[16]. The mere presence or absence of a variant in such control data sets is only partially informative, as determinationof the pathogenicity of inherited CNVs is at present limited
by a lack of information on CNV frequency and combinations in apparently healthy individuals. Based on these considerations, examinations of a relatively large number ofindividuals from various specific ethnic groups have recently
been conducted using diﬀerent array platforms, such as BAC
arrays [6, 17, 18], oligo arrays [19–21], and others [22]. The
results are not always consistent, and it is likely that diﬀerent
human populations bear diﬀerent inherited CNVs. The

2numbers of Japanese individuals examined to date are smaller than those in studies of other ethnicities [4]. Polymorphic
CNVs have received considerable attention since they mightplay an important role in the etiology of common diseases.
Therefore, more data regarding CNVs should be accumulated from Japanese populations. We summarized the available data by focusing on highly polymorphic CNVs (in≥5.0% of the individuals) in a previous paper [23]. However,rare variations have recently received attention from scientists who espouse a hypothesis called “common disease andrare variants,” in contrast to the “common disease-common
variants” hypothesis that underpins most genome-wide association studies. In this paper, we focus on CNVs observed atrelatively low frequency (<5.0% of the individuals) in a population residing in Hiroshima and Nagasaki, Japan, using
CGH with BAC clones as targets.

2. Materials
The study was conducted in two stages: Stage (1) 80 unrelated
Japanese individuals were examined by BAC-aCGH with anarray having 2,241 BAC clones; Stage (2) 133 unrelated Japanese individuals were examined by BAC-aCGH containing
2,622 BAC clones.
2.1. Target BAC Clones. The majority of the clones used inStage 1 of this study were selected from the set of cytogenetically mapped P1 artificial chromosome (PAC) clones andbacterial artificial chromosome (BAC) clones reported by the
BAC Resource Consortium [26] and obtained from either theChildren’s Hospital Oakland Research Institute (Oakland,
CA, USA) or from Invitrogen Inc., Co. (Carlsbad, CA, USA).
In Stage 2, together with the BAC clones in Stage 1, additional
381 BAC clones were used, a majority of which were collaboratively obtained from Dr. Matsumoto of Yokohama City
University. A total of 2,241 clones of chromosomal fragmentsfrom chromosome 1 to chromosome 22 were used in Stage 1
and 2,622 clones in Stage 2. The clones were distributed every
1.2 Mb across all human autosomes in Stage 1 and every 1.1
Mb in Stage 2. In addition to autosomal clones, four kindsof X-chromosomal clones were used as internal references.
With respect to the examinations in Stage 1, three sets ofarrays were constructed as described in the previous paper[27]. For Stage 2, all of the clones were printed onto one glass
slide.
2.2. Genomic DNA Samples. The genomic DNA samples
used in this study were reported in a previous study [23,27]. In brief, the DNA used for the population studies andthe family studies was extracted from lymphoblastoid celllines obtained from the oﬀspring of atomic-bomb survivors
and their family members. Lymphoblastoid cell lines were
derived from a cryopreserved archive of families consisting offather, mother, and oﬀspring from Hiroshima and Nagasaki
for whom permanent cell lines have been established by
Epstein-Barr (EB) virus transformation. The genomic DNA
was isolated using conventional methods as described indetail elsewhere [28]. A total of 305 of the oﬀsprings werescreened initially. Since the oﬀspring in some cases included

Journal of Biomedicine and Biotechnology
siblings, we selected one representative oﬀspring to create agroup of “unrelated individuals,” thereby avoiding any duplication of CNVs from families containing two or more siblings, in accordance with the rules described in a previous
report [23]. In this study, 80 individuals (40 from each city)were included in examinations in Stage 1 and 133 (84 from
Hiroshima, 49 from Nagasaki) in Stage 2. The individuals
provided their informed consent prior to the study. The Institutional Review Board of our foundation approved thisstudy.

3. Methods
3.1. Array Preparation. The arrays were prepared as described in the previous paper [23]. In brief, with respect toStage 1, cloned DNA was digested by NotI. On the other
hand, in Stage 2, cloned DNA was digested with MseI, andthe fragmented DNAs were amplified by ligation-mediated
PCR carried out as described by Snijders et al. [29]. In bothstages, the target DNAs (0.5 μg/μL) were dissolved in 50%
dimethylsuloxide and printed in triplicate onto the glass
slides (Matsunami Glass Co. Ltd.) using the Aﬀymetrix 417
Arrayer (Aﬀymetrix).
3.2. Screening by the Array CGH Method. The screeningsof both stages were conducted following the procedures
described previously [23]. In brief, for labeling DNA, test andreference genomic DNA (1.25 μg each) was cut by BamHI
and labeled by a random-priming method with Cyanine5- and Cyanine-3-labeled dUTP (Cy5- and Cy3-dUTP;
PerkinElmer Life Sciences, Wellesley, MA, USA). Human
CotI DNA was employed to block repetitive sequences of thelabeled probes.
Prehybridization was conducted to block repetitive sequence binding of target DNA on the arrays and to prevent
nonspecific binding of probe DNA to the targets. Following
the initial incubation, the prehybridization solution was removed, and hybridization solution with Cy-labeled DNA(prepared as described above) was added. The prehybridization and the hybridization were conducted with continuous
mixing overnight at 37◦ C. After hybridization, the arrays
were washed by the procedures reported previously [27]. Allof the procedures were conducted using the GeneTAC Hybridization Station (Genomic Solutions Inc., Ann Arbor, MI,USA).
After washing procedures, fluorescent images of the hybridized arrays were obtained using a ScanArray 5000 confocal
laser scanner (PerkinElmer Life Sciences). ArraySuite (Scanalytics Inc., Fairfax, VA, USA) in Stage 1 and Gene Pix (Axon
Instruments, Sunnyvale, CA) in Stage 2 were used to quantify
the fluorescence of each spot on the array images. We thenprocessed the data using software specifically developed to
identify “CNVs” [27].
3.3. Procedures of Population Studies. The population studies
were conducted in accordance with accepted procedures as
described in the previous paper [27]. In brief, hybridization
was performed as follows: mixtures of (a) Cy5-labeled reference DNA and Cy3-test DNA, and (b) inverse labeling with

Journal of Biomedicine and Biotechnology

3
(bp)

Chr15

44650000

44600000

44700000

44750000

Relative copy number

RP11-81H5

2

1

0
Oﬀspring
FatherMotherFigure 1: Characterization of a loss-type CNV detected on a BAC-clone RP11-81H5. A genome map of the RP11-81H5-containing region
was obtained from the UCSC Genome Browser website. The distance (bp) from the p-terminal of chromosome 15 is shown in the uppermost
part. The positions of PCR primers examined are shown by solid boxes. The experimental conditions are described more precisely in the text.
The relative copy number of each site was normalized using the quantity of the amplified segment of a portion of a diﬀerent chromosome(Chromosome 16) as a standard (relative copy number = 2) and was shown as mean ± SD (n = 3).

Cy3-reference DNA and Cy5-test DNA were applied to theslides. The two complementary hybridizations (i.e., (a) and(b)) were conducted for each individual. The variations ina given sample from a single individual that were identified
consistently in both complementary hybridizations ((a) and(b)) were regarded as putative CNVs. Since only DNA segments detected as putative CNVs in two individuals or more
were regarded as true CNVs, further analyses were not conducted. On the contrary, when CNVs were detected in only
one individual, all were confirmed with repeated examinations using arrays.
3.4. The qPCR Procedure for CNVs Identified in Oﬀspring.
The qPCR procedure was performed using SYBR premix EX
Taq (Takara-Bio, Ohtsu, Japan) and the Light Cycler System (Roche Diagnostics Japan, Tokyo, Japan), according to
the procedures described in the previous paper [27]. The
results were analyzed with Light Cycler Data Analysis software using a second derivative maximum model. Relative
DNA content of variants for every amplimer was calculatedusing a “normal” individual without CNVs as controls. The
relative copy number of each site was normalized using quantity of the amplified segment of a portion of a diﬀerent chromosome (chromosome 16:78888541–78888690; this position empirically showed the smallest deviation among many
samples in our experiments) as a standard (relative copy
number = 2) and was indicated as mean ± SD (n = 3).

3.5. Procedure of Family Studies for Confirmation of Inheritance of CNVs Identified Once in This Study. For the CNVs
identified in only one oﬀspring, further examinations werecarried out for confirmation of inheritance. The family studies using qPCR for the CNVs were conducted using DNAs
from the mother, the father and sibling of oﬀspring, if available. The procedures of qPCR were the same as above.

4. Results
4.1. Population Studies. We examined 213 unrelated individuals. The main purpose of this paper is to report the data
accumulated about CNVs found in <5.0% of the individuals.
(The number of CNVs was less than 10 in each BAC region.)
The results obtained from the population studies are shownin Tables 1 and 2. In Table 1, the CNVs identified in two
individuals or more are summarized. On the other hand, inTable 2, the CNVs identified in only one individual throughthis study are summarized. A total of 126 CNVs wereidentified at 52 diﬀerent BAC regions in the genome. The
CNVs observed at 27 of 52 BAC regions were found in only
one unrelated individual, and all of them were confirmedby repetitive analysis using the same array system as in the
population studies. The family studies by qPCR were conducted for 23 of 27 CNVs, and the results revealed that those
23 CNVs were inherited from one of the parents. The inheritance of CNVs from the remaining two oﬀsprings was not

4

Journal of Biomedicine and Biotechnology(bp)25950000

25900000

Chr8

26000000

RP11-90M13
PPP2R2A
EBF2

Relative copy number

3

2

1

0

6th intronof EBF2

Promoterregion of EBF2

Oﬀspring
Father

7th intronof PPP2R2A

MotherControl

Figure 2: Characterization of a loss-type CNV detected on a BAC-clone RP11-90M13. A genome map of the RP11-90M13-containing region
was obtained from the UCSC Genome Browser website. The distance (bp) from the p-terminal of chromosome 8 is shown in the uppermost
part. That region contains two genes, protein phosphatase 2, regulatory subunit B, alpha (PPP2R2A) and early B-cell factor 2 (EBF2). The
positions of PCR primers examined are shown by solid boxes. The individual has variant fragments with a deletion that contains a part of
the 7th intron of the PPP2R2A gene and the promoter region of the EBF2 gene.

confirmed, since DNA from the parents was not obtained.
On the other hand, the CNVs on two BAC clones (RP1188B18, RP11-86B6) were identified in only one unrelated
individual. In fact, however, they were observed in individuals who were not selected as “unrelated” individuals.
The numbers of individuals who expressed the CNVs aredescribed in the parentheses in Table 2. For that reason, we
did not conduct family studies for those CNVs.
4.2. The Typical Patterns of Three Family Studies of CNVs
on BAC Clones by qPCR Analysis. As typical examples offamily studies, the results of qPCR carried out for the
CNVs observed in three BAC-clone regions are describedin Figure 1 (RP11-81H5), Figure 2 (RP11-90M13), andFigure 3 (RP11-90O18 and RP11-90C3). For the first case,a single 180,264 bps deletion was observed in an oﬀspring.
The family study demonstrated that the CNV was inheritedfrom the mother. There is no gene in this BAC clone region.
For the second case, the BAC-clone (RP11-90M13) contains
two genes, protein phosphatase 2, regulatory subunit B, alpha(PPP2R2A) and early B-cell factor 2 (EBF2). The deletiontype CNV was confirmed by qPCR, and the CNV was foundto be inherited from the father. The deletion covered part of
intron 7 of PPP2R2A and the promoter region of EBF2. For
the last case, as shown in Figure 3, the CNV was covered by

two BAC clones (RP11-90O18 and RP11-90C3). This result
was revealed by qPCR, and the CNV was found to be inherited from the father. The CNV contained the series of zincfinger protein families (ZNF 107, 138, 273, 117, 92).
4.3. Comparing Our Data with Published Data. Many segmental duplications have already been summarized in public databases, such as the Database of Genomic Variants(DGV Database; http://projects.tcag.ca/variation/), UCSC
Human Genome Browser (UCSC Database. http://genome.ucsc.edu/index.html), and NCBI Map Viewer (NCBI Database; http://www.ncbi.nlm.nih.gov/mapview/map search.cgi).
The known CNVs have also been reported in the same databases. However, when the CNVs reported in the databasesor papers were too small to be detected by our BAC-aCGH
methods, we assumed that they were diﬀerent from our
CNVs. The CNV data obtained in our studies, including an
indication of the presence or absence of CNVs already reported in other databases, are summarized in Tables 1 and 2.
4.4. Comparing Our Data with the Data of Japanese Publishedby Other Groups. Compared to Caucasian populations, very
little information about CNVs in Japanese populations has
been systematically screened. We compared our data with thedata of Japanese populations from the HapMap project asJournal of Biomedicine and Biotechnology

5(bp)63800000

64000000

64200000

Chr7

64400000

200 Kb

RP11-90O18

RP11-90OC3

Relative copy number

ZNF107

ZNF138ZNF273

ZNF117

ZNF92

3
2
1
0
2nd intronof ZNF107
Oﬀspring
FatherMotherControl

Figure 3: Characterization of a gain-type CNV detected on a BAC-clone RP11-90O18 and RP11-90C3. A genome map of both RP11-90
O18- and RP11-90C3-containing regions was obtained from the UCSC Genome Browser website. The distance (bp) from the p-terminal ofchromosome 7 is shown in the uppermost part. That region contains zinc finger print protein gene families, that is, ZNF 107, 138, 273, 117,and 92. The positions of PCR primers examined are shown by the solid boxes.

reported by Redon et al. [4], in which CNVs were examinedby tiling BAC array. In addition to the data by BAC array,three groups, including Redon et al. [4], reported the dataof Japanese populations based on examinations conductedusing oligo-array methods, although each group used a different array platform [24, 25]. These results are also summarized in Tables 1 and 2. In the case where a multiple numberof CNVs were identified in our study (Table 1), the CNVs
in 16 of 25 BAC regions were reported in the other reports.
Moreover, regarding one individual, the CNVs identified innine of 27 BAC regions were reported in the data reportedby the others, although we should emphasize again that the
CNVs identified in our BAC region are not exactly the sameas those reported by the others. The majority of CNVs identified in our study tend not to be identified in the other reports. Thus, they can be considered novel CNVs in the general Japanese population.
4.5. Genes and Disease-Related Genes. We summarized someof the genes and disease-related genes that overlap with
the BAC-clone region for our CNVs (Table 3). In addition,
mRNAs also have been reported in the databases, but they
are too numerous to describe here. All BAC-clone regions
contained at least one mRNA, although the functions ofa majority of the mRNAs are not yet known (data notshown).

5. Discussion
We examined 213 unrelated Japanese individuals using BACaCGH and found a total of 126 CNVs on 52 diﬀerent BAC
regions in the genomes. The CNVs observed on 27 of the 52
BAC-regions were found in only one unrelated individual.
The family studies were conducted for 23 CNVs, and theresults demonstrated that these were inherited from one ofthe parents. A fraction of the regions involved in the CNVs
observed in our study (i.e., 34 of 126 or about 27%; Tables
1 and 2) were not reported previously in other studies listed
in the database. Thus, they are considered novel CNVs. In
contrast, the data of highly polymorphic CNVs [23] demonstrated that only 8% (55 of 680) had not been reported.
This result showed that a significantly large number of novel
CNVs have been identified among relatively less frequent
CNVs (χ 2 = 36.64, P = 5.10 × 10−10 ). This finding suggests that the structural rearrangements for creating less frequent CNVs are evolutionarily recent. We thus paid attention to diﬀerences in the number of CNVs identified in
Hiroshima compared to Nagasaki. With respect to the CNVs

6

Journal of Biomedicine and Biotechnology
Table 1: Summary of the CNVs detected in plural unrelated individuals.

Clone
number
RP11-81B15
RP11-91G12
RP11-118M12
RP11-91F17
RP11-101J14
RP11-80P12
RP11-23D23
RP11-90C3
RP11-35P20
RP11-45M12
RP11-113B7
RP11-89I12
RP11-79I19
RP11-310C13
RP11-79M9
RP11-326B4
RP11-89A14
RP11-531P20
RP11-143O10
RP11-91C17
RP11-194H7
RP11-152F13
RP11-88B2
RP11-13J15
CTA-526G4
Total of CNVs

Chrom. Position
number (Mb)
1
1
2
3
4
4
7
7
7
8
8
8
8
9
10
13
13
13
13
14
15
15
18
21
22

150.7
194.9
241.2
167.5
8.1
132.3
0.84
64.3
71.6
3.9
4.2
4.4
3.7
113.5
77.7
66.8
82.4
84.4
88.5
43.2
32.5
80.9
61.8
24.1
20.6

Redon(4) Redon(5)Known Segmental
Hiroshima(2) Nagasaki(3)variant duplication
Delete
[4]
[4]
(6)
(8)
(10)
0
Pre
Abs
4
5
Abs
Abs
0
Pre
Pre(9)
2
0
Abs
Abs
0
Pre
Pre
0
3
Pre(11)
Pre
0
Pre
Abs
2
0
Abs
Abs
0
Pre
Abs
0
2
Abs
Abs
0
Pre
Abs
1
1
Abs
Abs
0
Pre
Abs
3
1
Abs
Abs
1
Pre
Abs
1
1
Pre
Abs
0
Pre
Pre
0
2
Pre
Pre
2
Pre
Abs
2
0
Pre
Pre
4
Pre
Abs
4
0
Pre
Pre
4
Pre
Abs
4
0
Pre
Pre
4
Pre
Abs
4
0
Pre
Pre
1
Abs(7)
Abs
5
2
Abs
Abs
0
Abs
Abs
4
2
Abs
Abs
0
Abs
Abs
5
3
Abs
Abs
0
Pre
Abs
4
1
Abs
Abs
0
Pre
Abs
2
0
Abs
Pre
0
Pre
Abs
2
2
Abs
Abs
0
Abs
Abs
3
2
Abs
Abs
0
Pre
Pre
2
0
Pre
Abs
9
Pre
Pre
5
4
Pre
Abs
2
Pre
Abs
0
2
Pre
Abs
4
Pre
Abs
2
2
Abs
Abs
2
Pre
Abs
2
1
Pre
Abs
33
63
36

Variants
Amp(1)
9
2
3
2
2
2
4
1
2
0
0
0
0
6
6
8
5
2
4
5
2
0
0
0
1
66

Park
et al.
[24]
Pre
Pre
Abs
Abs
Abs
Abs
Abs
Pre
Pre
Pre
Abs
Pre
Abs
Abs
Abs
Abs
Abs
Abs
Pre
Abs
Abs
Abs
Pre
Abs
Abs

Ikeda
et al.
[25]
Abs
Pre
Abs
Abs
Abs
Abs
Abs
Pre
Abs
Abs
Abs
Abs
Abs
Abs
Abs
Abs
Abs
Abs
Abs
Abs
Abs
Abs
Abs
Pre
Pre

(1)

Amplification. Number of CNVs observed in Hiroshima individual (2) and in Nagasaki individual (3) . (4) Radon’s BAC data. (5) Radon’s oligo-array data. CNVs
present (6) or absent (7) in previous papers. Segmental duplication absent (8) or present (9) in BAC-clones on which CNVs were observed. CNVs absent (10) or
present (11) in Japanese.

observed multiple times, 12 of 25 were identified in one city.
These CNVs appear to be so-called “private polymorphisms”
occurring more recently than the others. On the other hand,quite a small number (21 of 126; about 17%) of the CNVs inthis report were found on the BAC clones that overlappedwith segmental duplication (Tables 1 and 2). A majority(about 63%) of highly polymorphic CNVs, however, wereobserved on the BAC-clone overlapping with segmental duplication (see Table 1 in [23]). This result suggests that segmental duplication might play a significant role in the creation of highly polymorphic CNVs. These observations aresupported by previous data from Sharp et al. [17], who reported the sharing of CNVs among several populations,meaning that the specific genomic imbalances either predated the migration of modern humans from Africa or arose
independently in diﬀerent populations.
On the other hand, the data representing Japanese populations are based on reports from three groups. We compared our CNV data to that of these other reports [4, 24, 25].
The CNVs found in 27 of 52 BAC regions (about 52%) were

not identified by the other groups. Speculation as to thereason for this includes the possibility that since the CNVs
were private polymorphisms as mentioned above, they were
not identified in the Japanese populations used in the otherworks.
In humans, it is estimated that around 23% of currently
known CNVs reside in known or putative genes. Many CNVs
that lie outside genic regions of the genome may still havea significant influence on gene expression by aﬀecting gene
regulatory elements [30]. Another important area of exploration is linkage disequilibrium (LD), that is, nonrandom
association between alleles at diﬀerent loci, between CNVs
and other genomic variants (especially SNPs), which may
provide important insight into the genomic evolution (e.g.,
the recombination patterns) of CNVs [31, 32]. However,studies of LD between CNVs and SNPs are hindered by
the fact that CNVs generally reside in genomic regions
associated with segmental duplications and/or repeat-richregions, which are diﬃcult to sequence and use for detectionof SNPs.

Journal of Biomedicine and Biotechnology

7

Table 2: Summary of the CNVs detected in single unrelated individuals.
Clone
number
RP5-895L2
RP11-204C23
RP11-80D22
RP11-22O8
RP11-104N13
RP11-90O18
RP11-79I19
RP11-90J21
RP11-90M13
RP11-91G3
RP11-91H19
RP11-176H12
RP11-18B9
RP11-75H24
RP11-90N7
RP11-86C3
RP11-80H14
RP11-81H5
RP11-90E5
RP11-79I15
RP11-89L6
CTB-15E16
RP11-265I6
RP11-89N1
RP5-908M14
Remarks
RP11-88B18
RP11-86B6

Chrom.
Position
DeleteKnown
Segmetal
Known
Redon Redon(3) Park et al. Ikeda et al.
[4]
[24]
[25]
number(Mb)
or Amp(1)variant duplication inheritance(2) [4]1
231.9
DeletionAbs(4)
Abs(6)MO
Abs(8)
Abs
Abs
Abs
(5)
3
3
Delete
Pre
Abs
FA
Abs
Abs
Abs
Abs
4
43.7
Amp
Pre
Abs
FA
Abs
Abs
Abs
Abs
4
141.1
Deletion
Abs
Abs
ND
Abs
Abs
Abs
Abs
6
165
Deletion
Pre
Abs
FA
Abs
Abs
Abs
Abs
7
63.8
Amp
Pre
Abs
FA
Abs
Abs
Abs
Abs
8
4
Delete
Pre
Abs
MO
Abs
Pre
Abs
Abs
8
8.8
Amp
Pre
Abs
FA
Abs
Abs
Abs
Abs
8
25.9
Deletion
Abs
Abs
FA
Abs
Abs
Abs
Abs
8
91.1
DeletionPre
Abs
ND
Pre(9)
Pre
Abs
Abs
10
58.2
Deletion
Pre
Abs
FA
Abs
Abs
Abs
Abs
10
65.7
Amp
Pre
Abs
MO
Abs
Abs
Abs
Abs
11
40.3
Delete
Pre
Abs
MO
Abs
Abs
Abs
Abs
(7)
11
58.6
Amp
Pre
Pre
MO
Pre
Abs
Pre
Abs
12
11.4
Amp
Pre
Pre
MO
Abs
Abs
Pre
Abs
13
89
Deletion
Pre
Abs
FA
Abs
Abs
Abs
Abs
15
20.5
Amp
Pre
Abs
FA
Abs
Abs
Abs
Pre
15
44.7
Delete
Abs
Abs
MO
Abs
Abs
Pre
Abs
15
98.4
Amp
Pre
Abs
FA
Abs
Abs
Abs
Abs
16
16.6
Deletion
Pre
Pre
MO
Pre
Pre
Abs
Abs
16
75.7
Amp
Abs
Abs
MO
Abs
Abs
Abs
Abs
16
79.0
Amp
Pre
Abs
MO
Abs
Abs
Abs
Abs
18
36.1
Delete
Abs
Abs
FA
Abs
Abs
Abs
Abs
18
76.0
Amp
Pre
Abs
MO
Abs
Abs
Abs
Pre
20
60.4
Amp
Pre
Abs
MO
Pre
Abs
Pre
Abs
Each CNV as below was detected in three individuals in diﬀerent families, but two of them were detected in individuals
who were not selected as an unrelated individual. Both CNVs were not identified in three other studies of Japanese
10
56.6
Deletion
Pre
Abs
ND
1 (3)
13
104
Amp
Abs
Abs
ND
1 (3)

(1)

Amplification. (2) CNVs were inherited from the mother (MO) or the father (FA). (3) Data from Aﬀymetrix oligo array. CNVs absent (4) or present (5) inprevious papers. Segmental duplication absent (6) or present (7) in BAC clones on which CNVs were observed. CNVs absent (8) or present (9) in Japanese datafrom three types of oligo arrays.

Recent association studies have attempted to link CNVs
with expression profiles, diseases, and known phenotypic
diﬀerences. On the other hand, distribution of CNVs diﬀers
depending on genetic background, and population-specific
CNVs might account for the divergence of some physiological traits and disease prevalence among populations [31].
Like SNPs, CNVs also show variability among populations;
therefore, comprehensive CNV information is needed when
applying CNVs as genetic markers in disease or trait studies.
For the reasons cited above, we made plans to accumulate
Japanese data in which we provide information about therelation between Japanese CNV data and phenotypic differences correlated with reported diseases. In our previous
paper [23], we summarized such data by focusing on highly
polymorphic CNVs found in ≥5.0% of the individuals.
More recent studies based on advanced molecular technologies, such as genome-wide association studies [33, 34]
and next generation sequencing [35, 36], reported that many

genes appear to play important roles in the etiology of common diseases. Depending on newly developed technologies,
rare variations have recently received attention from scientists who espouse a hypothesis called “common disease andrare variants.” In this paper, we summarized CNVs thatwere identified in fewer than 10 individuals in our previous
population study. We focused on genes reported in the
DGV, UCSC, and NCBI databases (Table 3), although many
mRNAs were also listed in them. Since these genes are goodcandidate markers for enabling us to examine the etiology ofcommon diseases and phenotypical heterogeneities amongindividuals, our CNVs have the potential to become usefulmarkers in future studies.

6. Conclusion
In this study, 34 CNVs were new, indicating that CNV coverage of the human genome is still incomplete and that there is

8

Journal of Biomedicine and Biotechnology
Table 3: Genes in BAC clones with CNVs.

BAC-clone number

Chrom. number

Position (Mb)

RP11-81B15

1

150.7

RP11-91G12

1

194.9

RP11-118M12

2

241.2

RP11-101J14

4

8.1

Genes
Late cornified envelope 5A (LCE5A), 3A(LCE3A), 3B(LCE3B), 3C(LCE3C),
3D(LCE3D), 3E(LCE3E), cysteine-rich C-terminal 1 (CRCT1)
Complement factor H (CFH). Complement factor H-related 3 (CFHR3)
Aquaporin 12B (AQP12B), aquaporin 12A (AQP12A), kinesin family
member 1A (KIF1A)
Actin binding LIM protein family, member 2 (ABLIM2)

RP11-23D23

7

0.84

ArfGAP with dual PH domains 1 (ADAP1)

RP11-90C3

7

64.3

Integrator complex subunit 4-like 1 (INTS4L1)

RP11-35P20

7

71.6

tRNA-yW synthesizing protein 1 homolog B (S.cerevisiae) (TYW1B)

RP11-45M128

3.9

CUB and Sushi multiple domains 1 (CSMD1)

RP11-113B78

4.2

CUB and Sushi multiple domains 1 (CSMD1)

RP11-89I128

4.4

CUB and Sushi multiple domains 1 (CSMD1)

RP11-79I19

8

3.7

CUB and Sushi multiple domains 1 (CSMD1)

RP11-310C13

9

113.5

Chromosome 9 open reading frame 84 (C9orf84)

RP11-79M9

10

77.7

Chromosome 10 open reading frame 11 (C10orf11)

RP11-326B4

13

66.8

Protocadherin 9 (PCDH9)

RP11-91C17

14

43.2

Rho GTPase activating protein 5 (ARHGAP5)

RP11-194H7

15

32.5

RP11-152F13

15

80.9

Golgin A8 family, member A (GOLGA8A)
Ubiquitin-conjugating enzyme E2Q family member 2 pseudogene 2(UBE2Q2P2), golgin A6 family-like 9 (GOLGA6L9), cytoplasmic
polyadenylation element binding protein 1, golgin A2 pseudogene(LOC727849), cytoplasmic polyadenylation element binding protein
1(CPEB1)

CTA-526G4

22

20.6

Topoisomerase (DNA) III beta (TOP3B)

RP11-90O18

7

63.8

Zinc finger protein 107 (ZNF107),zinc finger protein 138 (ZNF138)

RP11-79I19

8

4

CUB and Sushi multiple domains 1 (CSMD1)

RP11-90J21

8

8.8

Exoribonuclease 1 (ERI1)

RP11-90M13

8

25.9

Early B-cell factor 2 (EBF2)

RP11-91G3

8

91.1

RP11-90N7

12

11.4

RP11-80H14

15

20.5

Calbindin 1, 28 kDa (CALB1)
Proline-rich protein BstNI subfamily 4 (PRB4), proline-rich protein BstNI
subfamily 1 (PRB1), proline-rich protein BstNI subfamily 2 (PRB2)
Tubulin, gamma complex-associated protein 5 (TUBGCP5), cytoplasmic
FMR1 interacting protein 1 (CYFIP1)

RP11-81H5

15

44.7

RP11-90E5

15

98.4

RP11-89N1

18

76.0

RP5-908M14

20

60.4

MicroRNA 548a-3 (MIR548A3)ADAM metallopeptidase with thrombospondin type 1 motif, 17(ADAMTS17)
Ribosome binding factor A (putative) (RBFA), ADNP homeobox 2 (ADNP2),
hypothetical LOC100130522 (LOC100130522), par-6 partitioning defective 6
homolog gamma (C. elegans) (PARD6G)
Ribosomal protein S21 (RPS21), Cdk5 and Abl enzyme substrate 2(CABLES2), chromosome 20 open reading frame 151 (C20orf151)

diversity between the Japanese and other ethnic populations.
Moreover, the CNVs found in about two-thirds of the BAC
regions examined in our study were not identified in thestudies in which three groups examine Japanese populations
using diﬀerent array platforms. The newly identified CNVs
extend the coverage of CNVs in the human genome (also thegeneral Japanese population). Some CNVs contained genesthat might be related to phenotypic heterogeneity amongindividuals. Moreover, it is expected that the CNVs could
now be taken into consideration when genetic studies, forexample, CNV association studies, are conducted.

Authors’ Contribution
Y. Satoh and N. Takahashi equally contributed to this work as
first authors.

Journal of Biomedicine and Biotechnology

Acknowledgments
The authors thank H. Omine, J. Kaneko, A. Miura, M.
Imanaka, and E. Nishikori for their technical assistance. They
are grateful to E. B. Douple for critical reading of the paper.
This publication was supported by Research Protocols RP
1-01 and RP 2-07 of the Radiation Eﬀects Research Foundation (RERF) and in part by Grants-in-Aid for Scientific
Research from the Japanese Ministry of Education, Culture,
Sports, Science and Technology and the Japan Science andTechnology Agency (Core Research for Evolutional Scienceand Technology). RERF, Hiroshima and Nagasaki, Japan, isa private nonprofit foundation funded by the Japanese
Ministry of Health, Labour and Welfare and the U.S. Department of Energy, the latter in part through the National
Academy of Sciences.

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