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Genetic analysis of western North Pacific minke whales from Korea
and Japan based on microsatellite DNA
Naohisa Kanda1, Jung Youn Park2, Mutsuo Goto1, Yong-Rock An3, Seok-Gwan Choi3, Dae-Yeon Moon3,
Toshiya Kishiro4, Hideyoshi Yoshida4, Hidehiro Kato5, and Luis A. Pastene1
1

The Institute of Cetacean Research, Tokyo, Japan
Biotechnology Research Center, National Fisheries Research and Development Institute, Busan, Korea
3
Cetacean Research Institute, National Fisheries Research and Development Institute, Ulsan, Korea
4
National Research Institute of Far Seas Fisheries, Yokohama, Japan
5
Tokyo University of Marine Science and Technology, Tokyo, Japan
2

Contact email:kanda@cetacean.jp
ABSTRACTAt the final stage of the Implementation process for the western North Pacific (WNP) common minke
whales, the IWC Scientific Committee (SC) adopted four stock structure scenarios (baselines A, B, C, andD)(IWC, 2004). Previous papers by some of the current authors conducted stock identification at anindividual level using a Bayesian clustering method (Structure; Pritchard et al., 2000), and subsequently
examined the plausibility of these four stock baseline scenarios, based on the genetic variation at 16
microsatellite DNA markers analyzed from samples of minke whales collected during JARPNII as well as
JARPN from 1994 to 2007. The stock identification successfully distinguished most of the sampledwhales (91%) to the J and O stocks, and subsequent stock structure analysis with the identification
showed that 1) whales from the J stock existed in the SA7 west (7W) with low but large enough number
to cause genetic heterogeneity observed in the 7W samples as well as between the 7W and other samples,2) except the J stock whales, the survey area was mainly occupied by O stock, and 3) the stock structure
scenarios that assumed genetically distinct stock in the 7W (baselines C and D) were not supported. It
was argued, however, that Structure could have overlooked weakly differentiated stocks under the genetic
difference between the J and O stocks, and thus some, if not all, of the unassigned individuals from thestock identification may belong to different stocks from the J and O. Alternately, these individuals wereunassigned due to the lower statistical power than we have expected. In order to examine the status of
the unassigned individuals, this paper conducted some additional Structure runs and Principal Coordinate
analysis. The results of these analyses failed to detect evidence of the additional stocks and suggested
that these unassigned individuals were probably either J or O stocks. Similar to the previous papers, ourstudy support that WNP is mainly occupied by the O stock.
In addition to the stock structure in WNP, examination of the stock structure in Sea of Japan (SOJ)and Yellow Sea (YS) is important for pre Implementation assessment for common minke whales. The
SC listed seven stock structure hypotheses that proposed a single to maximum three stocks in SOJ and YS.
We combined the Korean genetic data of bycaught minke whales from 1999 to 2007 to the Japanesegenetic data to examine stock structure of them in the WNP, SOJ, and YS. The samples of 2005 to 2007
were not used in the previous papers. Both conventional hypothesis testing and Structure analysis wereconducted to identify the number of stocks in the SOJ and YS. The Structure analysis did not show
evidence of stock structure in SOJ and YS, while the conventional hypothesis testing detected very weak
genetic differences within the Korean samples as well as between the Korean and Japanese samples.
The results suggested there may be a sub-stock that mainly occupies YS (Y stock) and some whales in the
Y stock appear to migrate north along the Korean coast that could have caused the genetic difference
between the Korean and Japanese samples in SOJ. More samples from YS will be needed for further
analyses.
In summary, we accept the previous view that main stocks inhabiting the Korean and Japanesewaters were the J and O stock. In addition to the two stocks, the Y sub-stock may occupy YS but further
analyses with more samples will be needed to reach a final conclusion.
KEY WORDS: MINKE WHALE, MICROSATELLITE, STOCK STRUCTURE, NORTH PACIFIC, SEA
OF JAPAN, YELLOW SEA

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INTRODUCTION
Common minke whales, Balaenoptera acutorostrata, are the smallest and the most abundant baleen
whale species inhabiting major open oceans world-wide with spatial and temporal separations among
populations (Pastene et al., 2007). They live up to 50 years in age and the adult size is, on average,
6-7m. They feed on various prey species, such as copepods, Euphausiids, and fish. Their age at first
reproduction is five, and they are thought to reproduce every year. As typical baleen whales, minke
whales undergo seasonal movement from winter breeding grounds in low latitude to summer feeding
grounds in high latitude. Around the ocean off the Japanese coast, at least two different stocks of minke
whales are known to exist: one stock distributes in the western North Pacific (WNP) and the other in the
Sea of Japan (SOJ) (Omura and Sakiura, 1956; Ohsumi, 1977; Kato, 1992; Wada and Numachi, 1991;
Goto and Pastene, 1997; Pastene et al., 2007).
The IWC Scientific Committee (SC) completed the RMP Implementation for the WNP common minke
whales during the 2003 Annual Meeting. At the final stage of the Implementation process, the SC
adopted the following stock scenarios in the WNP (IWC, 2004).
(1) Baseline A: three-stock scenario (J, O, W) with the W stock found only in part of
sub-area 9 and only sporadically.
(2) Baseline B: two stock scenario (J and O) with no W stock as a limiting case of
Baseline A.
(3) Baseline C：four-stock scenario overall, with OW, OE and W to the east of Japan.
Boundaries are fixed at 147°E and 157°E and there is no mixing between the stocks.
(4) Baseline D：three-stock scenario (J, O, W), with O and W mixing over 147°E and162°E, O being dominant to the west and W to the east.
The SC did not examine the plausibility of each baseline scenario at all because it was afraid that any
conclusions would not have been accepted by all. Consequently, the SC rated all of the scenarios thesame ‘high’ plausibility.
In Kanda et al. (2009b; see also Kanda et al., 2009a and Goto et al., 2009), the plausibility of these fourbaseline stock scenarios was examined analyzing samples of minke whales collected from JARPN and
JARPNII conducted from 1994 to 2007 using 16 microsatellite DNA markers. Results of the paper
indicated 1) whales from the J stock existed in the 7W with low but large enough number to cause genetic
heterogeneity observed in the 7W samples as well as between the 7W and other samples, 2) except the J
stock whales, the survey area was mainly occupied by O stock, and 3) the baselines C and D were notsupported because no other genetically distinct stock was observed in the survey area.
Prior to conducting the analyses in Kanda et al. (2009b), Kanda et al. (2009a) genetically assigned eachindividual in the samples to either pure O, pure J and unassigned individuals using the Bayesianclustering method implemented in the computer program Structure (Pritchard et al. 2000). The tests forstock differentiation were then carried out with the suspected J stock individuals (all individuals included)
and without the suspected J stock individuals (individuals of unassigned and O stock included) as well aswith only the suspected O stock individuals (individuals of unassigned and the J stock excluded).
One of the arguments for the results of these papers was how to interpret the unassigned individuals.
Probably due to the low statistical power of the data, it was difficult to distinguish the two possibilities,
that is, they were from the J or O stocks or from additional stocks. Because the two stocks are notcompletely distinct, there may be individuals genetically indistinguishable to the J and O stocks.
Alternately, signals from the additional weakly differentiated stocks might have been overwhelmed by the
relatively large O-J differences.
In addition to the stock structure of minke whales in the North Pacific, Japanese and Korean scientists
have worked together to analyze the samples from Japan and Korea using mtDNA and microsatellites toinvestigate stock structure of common minke whales in SA5 and SA6 of the Japanese and Korean waters(Goto et al., 2007; Kanda et al., 2006; Park et al., 2006). On the basis of the results from these analyses,
the Working Group on the in-depth assessment of WNP common minke whales, with a focus on J stock,has listed following seven stock structure hypotheses for minke whales inhabiting Yellow Sea (YS), SOJ,
and Pacific coast of Japan (IWC, 2010).

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Do not cite without written permission from the authors(1) One stock, J: J stock migrates to YS, SOJ, and the Pacific coast of Japan.
(2) Two stocks, J and Y: J stock migrates along both coasts of Japan, and Y stock migrates along the
Korean coast.
(3) Two stocks, J and Y: J stock migrates through the SOJ and Pacific coast of Japan, and Y stock
migrates up to the YS.
(4) Two stocks, J and Y: J and Y stocks migrate through YS and SOJ at different times of year.
(5) Three stocks, J, K, and Y: J stock migrates along both coast of Japan, K stock along Korean coast, andY stock up to YS.
(6) Three stocks, Je, Jw and Y: Je stock migrates along the Pacific coast of Japan, Jw stock through SOJ,
and Y stock up to YS.
(7) Three stocks, Je, Jw and Y: Je stock migrates along the Pacific coast of Japan, Jw stock along the west
coast of Japan, and Y stock migrates along the Korean coast including YS.
Among the seven hypotheses, only the first four were agreed by the members of the working group.
The primary objectives of this study were two-fold. First, we attempted to better identify the status of
the unassigned individuals in the previous papers conducting analyses recommended from the working
group on the in-depth assessment of WNP common minke whales, with a focus on J-stock in the SC61
(IWC, 2009). We did some new Structure analyses with only unassigned individuals as well as withpure O plus unassigned individuals to detect the evidence of some additional, weakly-differentiatedstocks. Moreover, we also conducted Principal Coordinate analyses to better identify the status of theunassigned individuals to describe their relations in the two dimensional space.
Second, we combined the Korean genetic data of bycaught minke whales from 1999 to 2007 to theJapanese genetic data to examine stock structure of them in the WNP, SOJ, and YS. The samples of2005 to 2007 were not used in the previous papers. We conducted both conventional hypothesis testing
and Structure analysis to identify the number of stocks in the SOJ and YS.
MATERIALS AND METHODS
Samples
Table 1 shows the number of individuals used in this study for each of the sub-areas (SA) set for
management purpose of the WNP common minke whale (Fig.1). Common minke whale samples from1994-1999 JARPN and 2000-2007 JARPNII offshore component were used. These surveys wereconducted mainly summer in SA7, 8, 9, and 11. Another source of the minke whale samples was thecoastal component of the JARPNII survey conducted from 2002 to 2007. A total of nine surveys had
been conducted as the coastal component of the JARPNII: spring surveys at Sanriku in 2003, 2005, 2006,
and 2007, and fall surveys at Kushiro in 2002, 2004, 2005, 2006, and 2007. Minke whales that were
bycaught on set net fishery conducted along the Japanese coast from 2001 to 2007 were also used(bycatches). As of July 1st 2001, the new regulation governed by the Japanese government has allowed
the set net fishermen to harvest whales found in their set net and to sell these on to the market after DNA
registration of these for individual identification. The bycatches used were obtained from the SA2, SA6,
SA7, SA10, and SA11 year-round. Details about the samples can be found in Kanda et al. (2009a;
2009b), Kishiro et al. (2009), and Tamura et al. (2009).
Bycatches from Korea are those bycaught in coastal fishing gears along the Korean peninsula in SA6 andSA5 from 1999 to 2007 (Table 1). The Korean samples were divided into three on the basis of where
they came from: K6 = eastern side of the Korean Peninsula (SA6), K5-6 = southern side of the Korean
Peninsula (Tushima (Korean) Strait; SA5+SA6), and K5 = western side of the Korean Peninsula (SA5).
In order to look for evidence of seasonal variation for the bycatches, we first looked at the number of
bycatches by month from Korea (K6, K5-6, and K5 samples) and Japan (J6) (Table 2; Fig. 2). In SA6,
both the K6 and J6 showed a similar pattern that the number increased in spring to early summer as wellas in winter and decreased from summer to fall. Similar seasonal increase in May and decrease insummer was also observed in the K5-6 and K5 although the increase in winter was not observed. It
could be partially because of the small sample size. Based on these observations, we divided thesesamples from SA5 and 6 into three groups by their collection date to describe seasonal pattern of genetic
differentiation: MJ = March to June, JO = July to October, and NF = November to February.

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Microsatellite analysis
Skin tissues of minke whales taken during the JARPNII were stored in 95% ethanol until DNA extraction.
Genomic DNA was then extracted from 0.05g each of the skin tissues using standard proteinase K,
phenol-chloroform procedure described by Sambrook et al. (1989). Extracted DNA was stored in the
TE buffer (10 mM Tris-HCl, 1 mM EDTA, pH 8.0).
Microsatellite polymorphisms were analyzed using 16 sets of primers: EV1, EV14, EV21, EV37, EV94,(Valsecchi & Amos 1996), GT23, GT195, GT211, GT310, GT509, GT575 (Bérubé et al., 2000), GATA28,
GATA98, GATA417, TAA31 (Palsbøll et al., 1997), and DlrFCB14 (Buchanan et al., 1996). EV1, EV14,
EV21 were developed from sperm whale (Physeter macrocephalus), EV37, EV94, GT23, GT310, GT575,
GATA28, GATA98, GATA417, TAA31 were from humpback whale (Megaptera novaeanglia), and
DlrFCB14 from beluga whale (Delphinapterus leucas). All GT, EV, and DlrFCB primers were
dinucleotide repeat, TAA31 trinucleotide repeat, and all GATA primers tetranucleotide repeat. Primer
sequences and PCR profiles follows those of the original authors with slight modifications.
PCR amplifications were performed in 15µl reaction mixtures containing 10-100ng of DNA, 5 pmole ofeach primer, 0.625 units of Ex Taq DNA polymerase (Takara Shuzo), and 2mM of each dNTP, and 10x
reaction buffer containing 20mM MgCl2 (Takara Shuzo). PCR amplifications followed the
manufacture’s instructions for the use of Ex Taq DNA polymerase (Takara Shuzo). Amplified productswith internal size standard (GENESCAN400HD, Applied Biosystems Japan) were run on a 6%
polyacrylamide denaturating gel (Long RangerTM) using an BaseStationTM 100 DNA fragment analyzer(Bio-Rad). Although alleles were visualized using CartographerTM software specifically designed for
the BaseStation, allelic sizes were determined manually in relation to the internal size standard and minke
whale DNA of known size that were rerun on each gel.
Data analysis
The number of alleles per locus, allelic richness, and heterozygosity were computed using FSTAT 2.9.3(Goudet 1995). Conventional hypothesis testing procedure using heterogeneity test in frequencies of the
microsatellite alleles among samples was performed in the GENEPOP 4.0 (Rousset, 2008). Nullhypothesis to be tested is if the samples came from a genetically same group of minke whales. If genetic
differences exist, then it could indicate these samples came from genetically different stocks of minke
whales. Statistical significance was determined using the chi-square value obtained from summing thenegative logarithm of p-values over the 16 microsatellite loci (Sokal and Rohlf, 1995). FST was also
calculated using GENEPOP 4.0. The samples with less than 5 individuals were excluded from thegenetic divergence analyses.
The Bayesian clustering approach was implemented with the microsatellite data in the Structure 2.0(Pritchard et al., 2000) to determine the most likely number of genetically distinct stocks present in our
samples. Posterior probabilities for K were estimating from three independent runs for each value of K
from one to five with only genetic information. These data were calculated based on burn-in period of
10,000 iterations and runs of 100,000 iterations. Individual assignment was then conducted for the mostplausible K using estimated individual proportion of membership probability. The ancestry model weused for the simulation was the admixture model, which assumes individuals may have mixed ancestry.
The allele frequency model used was the correlated allele frequencies model, which assumes frequencies
in the different stocks are likely to be similar due to migration or shared ancestry.
Principal coordinate analysis (PCA) implemented in GenAlEx (Peakal et al., 2006) was used to describe
relationships among individuals with ordination. PCA is a multivariate technique that allows us to findand plot the major patterns within a multivariate data set (e.g., multiple loci and multiple samples).
GenAlEx is a computer program for population genetic analysis running within Microsoft ExcelTM. For
PCA, we needed to convert genetic data into a pairwise individual by individual genetic distance matrix.
The genetic distance matrices generated then form the input of PCA. Within the GenAlEx program, apairwise individual-by-individual (N x N) genotypic distance matrix is calculated for codominant datausing the following rules: For a single-locus, with i-th, j-th, k-th and l-th different alleles, a set of squared
distances is defined as d2(ii, ii) = 0, d2(ij, ij) = 0, d2(ii, ij) = 1, d2(ij, ik) = 1, d2(ij, kl) = 2, d2(ii, jk) = 3,
and d2(ii, jj) = 4. Genetic distances are summed across loci under the assumption of independence.
The matrix generated is then used for subsequent PCA analysis.

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DNA data quality control under the IWC guidelines
In order to minimize errors in genetic data, we were recommended to follow the IWC approved
guidelines for DNA data quality control for genetic studies (IWC, 2009). Since JARPN started, we haveestablished standard sampling, sample handling, and laboratory practice procedures to ensure consistent
data generation. These procedures are quite well consistent to the IWC guidelines except we did notscore microsatellite alleles by two persons for double-check. During the data collection, someindividuals in the samples were genotyped more than twice for double check. For data consistency andplausibility, we used the computer program MICRO-CHECKER (van Oosterhout et al., 2004) to checkfor null alleles and reading/typing errors. We also used the GENEPOP to test deviation from
Hardy-Weinberg equilibrium expectations and linkage disequilibrium in our samples. We conducted
these tests for several different sample groups (by each of sub-areas, SA7 to 9, by stocks (O, J, andunassigned individuals), all Korea, etc.) respectively, and found significant deviations within some groups
even after corrections for multiple tests (Rice, 1998) (data not shown). Because the loci involved in thedeviations and linkage disequilibrium were not consistent over the sample groups, however, we did notremove any ones from the 16 microsatellite loci for the further analyses. Finally, the guidelines
recommended to sequence the microsatellite loci for marker validation. Although we have not had theopportunity yet to do so by ourselves, we have used the published primers that were designed from thecetacean species and were checked for the reliability of amplification and scoring prior to use.
RESULTSFor genetic analyses of only the Japanese samples, we examined genetic variation at the 16 microsatellite
loci as described above. For genetic analyses of the Korean and Japanese samples combined, we
examined genetic variation at 11 of the 16 microsatellite loci because of difficulty of data standardization
between the Korean and Japanese labs.
Stock structure at the Pacific side of Japan (SA7-SA9)
Structure analyses with selected individuals in the samples
On the basis of the results from the individual identifications to stocks according to the criteria in Kanda
et al. (2009a), we conducted some new Structure runs focusing on the pure O and unassigned individuals
to detect some additional, weakly-differentiated stocks. We first conducted Structure with the samplesof the pure O and unassigned individuals (N=1772) combined (i.e., pure J excluded) and then with that of
only the unassigned individuals (N=240). Both cases presented the highest likelihood probability at
K=1 (Table 3). We were unable to detect signals of population differentiation.
Principal coordinate analysis on all samples
PCA was used to describe the relationships among the individuals from all the Japanese samples(N=2542) in a two dimensional space (Fig. 3). We plotted each individuals with the marks based on theindividual identification to the stocks (Kanda et al., 2009a) to see if there were any concordance between
the two independent analyses. The individuals assigned to the pure O and J stocks tended to occupy
different spaces, while most of the unassigned individuals tended to occupy the middle space of the pure
O and J. PC1 and PC2 explain 25.9 and 15.7 of variance, respectively.
Genetic difference between SOJ side and Pacific side of Japan (SA6 and SA2)
Genetic difference between the bycatch samples from SA6 and SA2 was examined using the genetic
variation at 16 microsatellite loci (Table 4). With all the individuals from the samples, the genetic
difference was observed between the two areas. However, after removing the pure O stock individuals,that difference was disappeared. Even for the former case, FST was quite low.
The observed genetic difference should have been resulted from the different proportion of the J and O
stock existed in these areas. Approximately 13% of the individuals in the SA2 sample (24/183) wereassigned to the O stock compared to only 1% in the SA6 sample (4/411).
Stock structure in SOJ and YS (SA5 and SA6)
Table 5 shows some genetic indices observed from the Korean amd Japanese samples at the 11
microsatellite loci. For the Japanese sample, although we demonstrated no structuring between SA2 andSA6, we used only the bycathes from SA6 to avoid the effect of the pure O individuals in the SA2 sample
that was difficult to remove completely. The numbers of alleles per locus and allelic richness were5

SC/62/NPM11
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were similar to each other.
Genetic differentiation between seasonal groups within same areas
We first looked for evidence of genetic difference at the 11 microsatellite loci among the seasonal groups
within the same area (Table 6). In the Korean samples, although no significant difference was detectedin the K5-6 and K5, significant genetic difference was observed among the groups in the K6. Pair-wisecomparisons indicated that the observed difference was due to the JO group. We thus combined the MJ
and NF groups into one as K6-MF and separated from K6-JO in the K6. No such difference wasobserved in the K5-6 and K5 although it could be due to the small sample size. In the J6, no suchseasonal difference was detected even though the samples size was large. In these three samples, threeseasonal groups in the K5-6, K5, and J6 were combined into one, respectively, as K5-6, K5, and J6.
Genetic differentiation among the areas
We looked for evidence of genetic difference among the K6-MF, K6-JO, K5-6, K5, and J6 (Table 7).
Within the Korean samples, the K6-MF showed significant difference to the K6-JO and K5, but not to theK5-6. The K6-JO did not show significant difference to the K5-6 and K5. The K5-6 did not show
significant difference to the K5. FST values among these comparisons ranged from 0.0004 to 0.0044.
The J6 showed significant differences to all of the Korean samples, and FST values ranged from 0.0004 to
0.0042. The lowest FST value was observed at the comparison between the J6 and K6-MF, which wasmuch lower than that between the K6-MF and K6-JO.
Although statistically significant differences were detected among the Korean and Japanese samples fromthe heterogeneity tests, Structure analysis did not support the existence of multiple stocks in the area
(Table 8). PCA was also used to describe the relationships among the individuals from the SA5 and SA6(Fig. 4). All of the individuals gathered in the same space. PC1 and PC2 explain 19.4 and 19.0 of
variance, respectively.
Stock structure in WNP, SOJ, and YSStructure analysis for all of the available samples was conducted with the genetic data from the 11
microsatellites. Structure with the total samples (3019 individuals) without information on theirgeographic origins presented the highest likelihood probability at K=2 (Table 9). Similar to Kanda et al.
(2009a), we assigned the individuals into the different groups on the basis of their membershipprobabilities over 0.900. All other individuals with the membership probability less than 0.900 to theeither groups were assigned as individuals of unknown origin (unassigned individual). With thiscriterion, 2441 individuals (81%) were assigned to the either stocks (1085 to stock 1 and 1356 to stock 2).
The proportion of the individuals with the membership probabilities over 0.900 was less than thatobserved from the analysis conducted in Kanda et al. (2009) using only the Japanese samples with thegenetic data at the 16 microsatellites. This lower proportion could have been due to lower number ofmicrosatellite loci used. Again, Structure was not able to detect the genetic difference within the Korean
and Japanese samples we detected using the hypothesis testing.
Spatial distribution of the assigned individuals
Both of the assigned and unassigned individuals were grouped based on their sampling origins (offshore,coastal, bycatch, Korea, and Japan) as well as locations (IWC sub-areas) (Table 10; Fig. 5). Similar tothe previous study (Kanda et al., 2009), distribution of the pure individuals that were genetically assigned
to the different stock was clearly separated geographically. Most of the individuals collected from the
SOJ and YS belonged to the Stock 1, whereas almost all of the individuals from the offshore NP (east of
the SA7E) belonged to the stock 2. Intermediate areas (SA7W and SA11) contained the individuals
from the both stocks. The results again indicated that the Stock 1 and Stock 2 were the J and O stock.
Unassigned individuals also distributed all areas with similar proportions.
DISCUSSION
We believe that the results of this study substantially improve our knowledge of the stock structure ofminke whales in the WNP, SOJ, and YS.
In the first part of this study, we focused on the unassigned individuals that were not assigned to either the
J or O stocks from the previous Structure analysis (Kanda et al., 2009a). The unassigned individuals

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could be the whales form some other additional weakly differentiated stocks or could be the whales thatwere not assigned to the J or O stock due to the low statistical power of the current genetic data. Bothadditional Structure runs and PCA analysis failed to detect the evidence of additional stocks in the area.
In the PCA analysis, although the J and O stock individuals tended to occupy the different spaces, theunassigned individuals tended to occupy the inbetween space, suggesting the unassigned individuals were
either the J or O stocks but not the additional stock. Considering the level of the genetic differences
between the J and O stocks (Kanda et al., 2009a), it is unlikely that all of these individuals were thehybrids between the two stocks. Addition of more number of microsatellite loci may better identify thestatus of the unassigned individuals.
We found the seasonal differences in the bycatch numbers in the Korean and Japanese samples from SA6.
The number of bycatches increased in winter and spring but decreased in summer. Similar, but not asclear, pattern was observed in the Korean samples from SA5. Although the similar seasonal pattern wasseen in both the Korean and Japanese samples, evidence of genetic difference between the seasonalgroups was only detected in the Korean samples (roughly, winter/spring group x summer/fall group).
Furthermore, only one of the two groups (the spring/winter group) from SA6 differed from the samples
from SA5. Considering the resident Y sub-stock in YS described in the stock structure hypotheses (IWC,2010), the evidence of genetic differences we found in the Korean samples might have been due to thestock difference between the J stock and Y sub-stock. We also found the yearly heterogeneity in thewinter group (p<0.05). This yearly heterogeneity and the very weak heterogeneity between the Japaneseand Korean samples from SA6 could be because individuals in the Y sub-stock might migrate to north atsome extent along the Korean coast. Each of the SA6 Korean year samples might contain theindividuals from the Y stock in different proportions. However, it is important to note that the recent
mitochondrial DNA analysis indicated no evidence of sub-stocks in the Korean minke whales (Park et al.,2009). Park et al. (2009) analyzed the Korean bycatch samples from 1998 to 2005, so that most of theindividuals were the same ones to this study.
The most likely explanation for the results of this study is that the main stocks inhabiting WNP, SOJ, andYS are the J and O stock, and that there maybe the Y sub-stock in YS. Further analyses with more
samples and more markers will be needed to reach a final conclusion especially for the Y sub-stock.
ACKNOWLEDGEMENTS
We thank researchers and crewmembers participating in the JARPN and JARPNII for their effort incollecting the samples used in this study. Our sincere gratitude also goes to H. Oikawa and S. Azumi for
their assistance in DNA extraction, and H. Hatanaka for valuable comments on the paper.
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8

SC/62/NPM11
Do not cite without written permission from the authors
Table 1. Number of individuals from the Korean and Japanese samples used in this study.
Total SA2 SA5 SA5+6 SA6 SA7W SA7E SA8W SA8E SA9W SA9E SA10 SA11Japan
JARPN & JARPNII
Offshore (94-07) 1231
414 47 86 139 291 174
80
Coastal (02-07)480
480
Bycatch (01-07)
831 183
411 212
9 16
Korea (99-07)

477

42

56 379

Table 2. Number of bycatches per month in SA5 and SA6 in Korea (99-07) and Japan(01-07).
Jan. Feb. Mar. Apr. May Jun. Jul. Aug. Sep. Oct. Nov. Dec. Total
Korea
SA6 39
20
31
36
39
43
17
18
26
22
36
52
379
SA5+6 4
1
2
5
15
13
6
3
0
0
5
2
56
SA5
4
1
2
0
4
12
4
4
3
2
3
3
42
Total

47

22

35

41

58

68

27

25

29

24

44

57

477

Japan
SA6

61

35

38

60

49

26

16

24

13

14

31

44

411

Table3. Results of Bayesian clustering method analyzed for the Japanese samples from SA7-SA9.
K
Log P(k/x)*
variance*Pr(k/x)**O and unassigned
1-90909.8
88.7
~1.0
2
-91939.7
2527.1~0.0
3-93413.2
5532.1~0.0
4-101177.4
21271.3~0.0
5-100444.1
19969.6~0.0
Only unassigned
1-11677.1
68.1
~1.0
2
-11713.5
212.2
~0.0
3
-11882.3
564.8
~0.0
4
-12162.2
1131.5~0.0
5-11937.4633.2~0.0* log likelihood of the data for different values of K and the variance. ** Probability for each of K.

Table 4. Results of the heterogeneity tests between the Japanese bycatches from SA2 and SA6.
were conducted with all individuals and without the O stock individuals.
SA2 x SA6
chi-square
FST
All individuals*
0.0008
Without O
ns-0.0003* = statistically significant, ns = no significant

9

Tests

SC/62/NPM11
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Table 5. The number of alleles (A), allelic richness (AR), and expected heterozygosity (He) at 11
microsatellite loci analyzed in the samples of minke whales used in this study.
Korea all (N=477)
Japan SA6 (411)
A
AR
He
A
AR
He
EV37
14 13.6 0.673
7
7.0 0.652
EV1
21 20.4 0.697
18 18.0 0.671
GT575
13 12.6 0.750
10 10.0 0.735
EV947
7.0 0.581
4
4.0 0.568
GT23
14 13.6 0.873
12 12.0 0.875
GATA988
7.7 0.590
5
5.0 0.568
GATA4179
8.7 0.670
8
8.0 0.673
GT211
13 12.6 0.775
12 12.0 0.803
EV214
4.0 0.334
2
2.0 0.344
DlrFCB145
4.9 0.413
3
3.0 0.452
GT195
15 14.8 0.812
8
8.0 0.802
Average11.2 10.9 0.652

8.1

8.1 0.649

Table 6. Results of the heterogeneity tests between the seasonal groups in the Korean and Japanesesamples from SA5 and SA6.
all
MJ x JO MJ x NF JO x NF
Korea SA6**
ns*
Korea SA5-6
ns------Korea SA5
ns------Japan SA6
ns------MJ = March to June, JO = July to October, NF = November to February

Table 7. Resutls of the pairwise heterogeneity tests and FST among the Korean and Japanese samples
from SA5 and SA6.
chi-square FST
Korea SA6 NJ
xKorea SA6 JO*
0.0029Korea SA6 NJ
xKorea SA5-6
ns
0.0008
Korea SA6 NJ
xKorea SA5*
0.0044
Korea SA6 NJ
x
Japan SA6*
0.0004
Korea SA6 JOx
Korea SA5-6
ns0.0015
Korea SA6 JOx
Korea SA5
ns
0.0044
Korea SA6 JOx
Japan SA6*
0.0042
Korea SA5-6
x
Korea SA5ns
0.0004
Korea SA5-6
x
Japan SA6*
0.0015
Korea SA5x
Japan SA6*
0.0039NJ = November to June; JO = July to October; * = statistically significant; ns = no significant

Table 8. Results of Structure runs for the Korean and Japanese samples from SA5 and SA6.
K
Log P(k/x)*
variance*Pr(k/x)**1-26839.4
57.2
~1.0
2
-27136.4
1043.4~0.0
3-27544.5
2176.0~0.0
4-27679.5
2787.1~0.0
5-28101.3
3886.6~0.0* log likelihood of the data for different values of K and the variance. ** Probability for each of K.

10

SC/62/NPM11
Do not cite without written permission from the authors
Table 9. Results of Structure runs for all of the Korean and Japanese samples (11 loci).
K
Log P(k/x)*
variance*Pr(k/x)**1-104341.4
72.6
~0.0
2
-100408.9
843.1
~1.0
3
-100977.6
2652.5~0.0
4-100723.1
3063.1~0.0
5-101459.8
4968.2~0.0* log likelihood of the data for different values of K and the variance. ** Probability for each of K.

Table 10. Structure analysis (11 loci) for all the samples from Japana and Korea samples
J stock
unassigned
O stock
Sum
Korea SA6
303
72
4
379
Korea SA5-6
44
11
1
56
Korea SA534
5
342
Japan SA10
7
2
0
9
Japan SA6
345
63
3
411
Japan SA2
130
33
20
183
Japan SA11
19
18
43
80
Japan SA11-BC
12
4
0
16
Japan SA7W-BC
92
45
75
212
Japan SA7W-K
34
51
168
253
Japan SA7W-S
33
55
139
227
Japan SA7W
25
76
312
413
Japan SA7E
0
6
42
48
Japan SA8W
1
18
67
86
Japan SA8E
0
31
108
139
Japan SA9W
6
55
230
291
Japan SA9E
0
33
141
174
Sum
proportion

1085
0.359

578
0.191

1356
0.449

3019

Fig. 1. Eighteen sub-areas used for the Implementation Simulation Trials for the North Pacific minke
whales.

11

SC/62/NPM11
Do not cite without written permission from the authors

70
Korea SA6
Korea SA5-6

60

Korea SA5
Japan SA6

50

40

30

20

10

0
Mar

Apr

May

Jun

Jul

Aug

Sep

Oct

Nov

Dec

Jan

Feb

Fig. 2. Number of bycatch individuals from SA5 and SA6 in Korea and Japan.

0.300
J stock
unassigned
O stock-0.400

0.000
0.000

0.400

-0.300Fig. 3. Results of the principal coordinate analysis (first two axes) among the individuals from all of theJapanese samples in the WNP. Assignment of the individuals was based on the Structure analyses
conducted with these individuals. See details in the text.

12SC/62/NPM11
Do not cite without written permission from the authors
0.450
Korea SA6
Korea SA56
Korea SA5
Japan SA6

0.000
0.000

-0.4500.450-0.450Fig. 4. Results of the principal coordinate analysis (first two axis) among the individuals from SA5 andSA6 in Korea and Japan.

J stock

unassigned

O stock

100%

80%

60%

40%

20%

0%
Korea Korea Korea
SA6 SA5-6 SA5

Japan
SA10

Japan
SA6

Japan
SA2

Japan Japan Japan Japan Japan Japan Japan Japan Japan Japan Japan
SA11 SA11- SA7W- SA7W- SA7W- SA7W SA7E SA8W SA8E SA9W SA9E
S
K
BC
BC

Fig. 5. Spatial distribution of the whales assigned to the Stock J, Stock O, and unassigned categories.
BC = bycatches, K = coastal survey of JARPNII at Kushiro, S = coastal survey of JARPNII at Sanriku.

13