Jang Wook Lee

Population genetic structure of Korean pen shell ( Atrina pectinata ) in Korea inferred from microsatellite marker analysis

The Korean pen shell Atrina pectinata is a commercially valuable species in Korea. The commercial catch of this fish has decreased continuously since 1990. However, its genetic characteristics have never been studied. In order to explore the population genetic structure of this species, 125 pen shells sampled from three major habitats along the western coast of Korea were genotyped at 21 microsatellite loci. Relatively high levels of genetic variability (mean number of allelic richness (AR) = 14.74; mean hetrerozygosity (He) = 0.849) was found among localities. However, despite the potential of the Korean pen shell to exhibit genetic structure (adults are sedentary), none of the genetic tests applied in this study detected significant genetic differentiation among the samples. The lack of genetic differentiation among samples may be due to high levels of larval dispersal, through passive drift with ocean currents. Alternatively, populations may have diverged too recently for significant genetic differentiation to have become evident. Furthermore, small sample sizes and the limited number of samples may have hampered the detection of genetic structure. The results of this study therefore suggested a lack of genetic structure among the populations of pen shell of Korean waters and should be managed as a single unit. This information on the genetic characteristics of Korean pen shell populations has important implications for the sustainable exploitation of the fishing resources and the preservation of biodiversity.

Wild and Hatchery Populations of Korean Starry Flounder (Platichthys stellatus) Compared Using Microsatellite DNA Markers

Starry flounder (Platichthys stellatus) is an important sport and food fish found around the margins of the North Pacific. Aquaculture production of this species in Korea has increased because of its commercial value. Microsatellite DNA markers are a useful DNA-based tool for monitoring the genetic variation of starry flounder populations. In this study, 12 polymorphic microsatellite DNA markers were identified from a partial genomic starry flounder DNA library enriched in CA repeats, and used to compare allelic variation between wild and hatchery starry flounder populations in Korea. All loci were readily amplified and demonstrated high allelic diversity, with the number of alleles ranging from 6 to 18 in the wild population and from 2 to 12 in the farmed population. A total of 136 alleles were detected at the 12 microsatellite loci in the two populations. The mean observed and expected heterozygosities were 0.62 and 0.68, respectively, in the hatchery samples and 0.67 and 0.75, respectively, in the wild samples. These results indicate lower genetic variability in the hatchery population as compared to the wild population. Significant shifts in allelic frequencies were detected at eight loci, which resulted in a small but significant genetic differences between the wild and hatchery populations (FST = 0.043, P < 0.05). Further studies with additional starry flounder sample collections are needed for comprehensive determinations of the genetic varieties between the wild and hatchery populations. These microsatellite loci may be valuable for future population genetic studies, monitoring the genetic variation for successful aquaculture management and the preservation of aquatic biodiversity.

Genetic Characterization of Five Hatchery Populations of the Pacific Abalone (Haliotis discus hannai) Using Microsatellite Markers

The Pacific abalone, Haliotis discus hannai, is a popular food in Eastern Asia. Aquacultural production of this species has increased because of recent resource declines, the growing consumption, and ongoing government-operated stock release programs. Therefore, the genetic characterization of hatchery populations is necessary to maintain the genetic diversity of this species and to develop more effective aquaculture practices. We analyzed the genetic structures of five cultured populations in Korea using six microsatellite markers. The number of alleles per locus ranged from 15 to 64, with an average of 23.5. The mean observed and expected heterozygosities were 0.797 and 0.904, respectively. The inbreeding coefficient FIS ranged from 0.054 to 0.184 (mean FIS = 0.121 ± 0.056). The genetic differentiation across all populations was low but significant (overall FST = 0.009, P < 0.01). Pairwise multilocus FST tests, estimates of genetic distance, and phylogenetic and principal component analyses did not show a consistent relationship between geographic and genetic distances. These results could reflect extensive aquaculture, the exchange of breeds and eggs between hatcheries and/or genetic drift due to intensive breeding practices. Thus, for optimal resource management, the genetic variation of hatchery stocks should be monitored and inbreeding controlled within the abalone stocks that are being released every year. This genetic information will be useful for the management of both H. discus hannai fisheries and the aquaculture industry.

Application of Novel Polymorphic Microsatellite Loci Identified in the Korean Pacific Abalone (Haliotis diversicolor supertexta (Haliotidae)) in the Genetic Characterization of Wild and Released Populations

The small abalone, Haliotis diversicolor supertexta, of the family Haliotidae, is one of the most important species of marine shellfish in eastern Asia. Over the past few decades, this species has drastically declined in Korea. Thus, hatchery-bred seeds have been released into natural coastal areas to compensate for the reduced fishery resources. However, information on the genetic background of the small abalone is scarce. In this study, 20 polymorphic microsatellite DNA markers were identified using next-generation sequencing techniques and used to compare allelic variation between wild and released abalone populations in Korea. Using high-throughput genomic sequencing, a total of 1516 (2.26%; average length of 385 bp) reads containing simple sequence repeats were obtained from 86,011 raw reads. Among the 99 loci screened, 28 amplified successfully, and 20 were polymorphic. When comparing allelic variation between wild and released abalone populations, a total of 243 different alleles were observed, with 18.7 alleles per locus. High genetic diversity (mean heterozygosity = 0.81; mean allelic number = 15.5) was observed in both populations. A statistical analysis of the fixation index (FST) and analysis of molecular variance (AMOVA) indicated limited genetic differences between the two populations (FST = 0.002, p > 0.05). Although no significant reductions in the genetic diversity were found in the released population compared with the wild population (p > 0.05), the genetic diversity parameters revealed that the seeds released for stock abundance had a different genetic composition. These differences are likely a result of hatchery selection and inbreeding. Additionally, all the primer pair sets were effectively amplified in another congeneric species, H. diversicolor diversicolor, indicating that these primers are useful for both abalone species. These microsatellite loci may be valuable for future aquaculture and population genetic studies aimed at developing conservation and management plans for these two abalone species.

Development of Microsatellite Markers for the Korean Mussel, Mytilus coruscus (Mytilidae) Using Next-Generation Sequencing

Mytilus coruscus (family Mytilidae) is one of the most important marine shellfish species in Korea. During the past few decades, this species has become endangered due to the loss of habitats and overfishing. Despite this species’ importance, information on its genetic background is scarce. In this study, we developed microsatellite markers for M. coruscus using next-generation sequencing. A total of 263,900 raw reads were obtained from a quarter-plate run on the 454 GS-FLX titanium platform, and 176,327 unique sequences were generated with an average length of 381 bp; 2569 (1.45%) sequences contained a minimum of five di- to tetra-nucleotide repeat motifs. Of the 51 loci screened, 46 were amplified successfully, and 22 were polymorphic among 30 individuals, with seven of trinucleotide repeats and three of tetranucleotide repeats. All loci exhibited high genetic variability, with an average of 17.32 alleles per locus, and the mean observed and expected heterozygosities were 0.67 and 0.90, respectively. In addition, cross-amplification was tested for all 22 loci in another congener species, M. galloprovincialis. None of the primer pairs resulted in effective amplification, which might be due to their high mutation rates. Our work demonstrated the utility of next-generation 454 sequencing as a method for the rapid and cost-effective identification of microsatellites. The high degree of polymorphism exhibited by the 22 newly developed microsatellites will be useful in future conservation genetic studies of this species.

Comparison between Wild and Hatchery Populations of Korean Pen Shell (Atrina pectinata) Using Microsatellite DNA Markers

Pen shell (Atrina pectinata) is a popular food source with a high commercial value in a number of Asian Pacific areas. The natural A. pectinata population has been declining continuously over the past several decades. Microsatellite DNA markers are a useful DNA-based tool for monitoring the genetic variation of pen shell populations. In this study, 20 polymorphic microsatellite (MS) DNA markers were identified from a partial genomic pen shell DNA library enriched in CA repeats, and used to compare allelic variation between wild and hatchery pen shell populations in Korea. A total of 438 alleles were detected at the 20 MS loci in the two populations. All loci were easily amplified and demonstrated allelic variability, with the number of alleles ranging from 5 to 35 in the wild population and from 5 to 22 in the farmed population. The average observed and expected heterozygosities were 0.69 and 0.82, respectively, in the hatchery samples and 0.69 and 0.83, respectively, in the wild samples. Statistical analysis of fixation index (FST) and analysis of molecular variance (AMOVA) showed minor, but significant, genetic differences between the wild and hatchery populations (FST = 0.0106, CI95% = 0.003–0.017). These microsatellite loci may be valuable for future aquaculture and population genetic studies for developing conservation and management plans. Further studies with additional pen shell samples are needed to conclusively determine the genetic diversity between the wild and hatchery populations.

Development and characterization of microsatellite markers for an endangered species, Epinephelus bruneus, to establish a conservation program

Kelp grouper (Epinephelus bruneus Bloch 1793) is a commercially important fish in Korea. In recent years, the catch of kelp grouper in the coastal waters of Korea has significantly declined. Despite its importance, little is known about its genetic diversity and conservation efforts are hampered. In this study, we isolated and characterized 12 microsatellite loci using an enrichment method based on magnetic/biotin capture of microsatellite sequences from a size-selected genomic library. All loci were readily amplified and contained TG/CA denucleotide repeats. To characterize each locus, 30 individuals from a natural E. bruneus population in the coastal waters of Jeju Island, Korea, were genotyped. All loci except three, KEm118, KEm154, and KEm219, were polymorphic, with an average of 8.1 alleles per locus (range 2–18). The mean observed and expected heterozygosities were 0.47 (range 0.19–1.00) and 0.61 (range 0.29–0.92), respectively. A significant deviation from Hardy-Weinberg equilibrium was observed at three loci (KEm134, KEm184, and KEm283). These findings will be useful for effective monitoring and management of genetic variation of kelp grouper as well as for the implementation of a fisheries conservation program.

Novel Polymorphic Microsatellite Loci for the Korean Black Scraper (Thamnaconus modestus), and Their Application to the Genetic Characterization of Wild and Farmed Populations

In this study, we developed 20 polymorphic microsatellite markers for the Korean black scraper, Thamnaconus modestus (Günther, 1877), Monacanthidae, and used them to compare allelic variation between wild and hatchery populations in Korea. All loci were readily amplified and demonstrated allelic variability, with the number of alleles ranging from 5–35 in the wild population and 5–22 in the farmed population. The average observed and expected heterozygosities were estimated, respectively, as 0.74 and 0.80 in the hatchery samples and 0.78 and 0.81 in the wild ones. These results indicate lower genetic variability in the hatchery population than in the wild population and minor, but significant, genetic differentiation between the two populations (FST = 0.005, P < 0.01). Additionally, cross-amplification was tested in another monacanthid species, Stephanolepis cirrhifer; many loci were found that yielded useful information. The high degree of polymorphism exhibited by the 20 microsatellites will be useful in future aquaculture and population genetic studies for developing conservation and management plans.

Molecular identification of Korean catfish (Siluriformes) based on two genetic markers

Eleven species of catfish (Siluriformes) found in Korean freshwater are economically important resources and include five endemic species. However, there are no studies on phylogenetic analysis of all catfish species in Korea at a molecular level. The species-level analysis of catfish species is usually carried out through morphological characters and controversial due to phenotypic variation. In this study, the partial sequences of 16S rRNA and cytochrome b mitochondrial genes were analyzed for species identification and phylogenetic relationships among 11 species of catfish from 10 different rivers in Korea. The nucleotide sequences of 16S rRNA and cytochrome b consisted of 587 and 441 nucleotide base pairs, respectively. Sequence analysis of both genes revealed that the 11 species fell into three distinct groups, which were genetically distinct from each other and exhibited identical phylogenetic resolution. Sequence divergences between congeneric species averaged 1.78% and 7.39% for 16S rRNA and cytochrome b, respectively. The phylogenetic relationships forming well-differentiated clades in the NJ, ML and BI trees were identical for both fragments. This research demonstrates that partial sequences of both the genes can efficiently identify the 11 species of catfish in Korea, indicating the usefulness of mtDNA-based approach in species identification.

New polymorphic microsatellite markers in the Korean mi-iuy croaker, Miichthys miiuy, and their application to the genetic characterization of wild and farmed populations

Eighteen new polymorphic microsatellite markers were developed for the Korean mi-iuy croaker (Miichthys miiuy, Perciformes, Sciaenidae), and allelic variability was compared between a wild population in Mokpo, Korea, and a hatchery population in Tongyeong, Korea. All loci were amplified readily and demonstrated allelic variability, with the number of alleles ranging from 5 to 37 in the wild population, and from 4 to 12 in the farmed population. The average observed and expected heterozygosities were estimated, respectively, to be 0.74 and 0.78 in the hatchery population samples, and 0.79 and 0.86 in the wild samples. These results indicate lower genetic variability in the hatchery population compared with the wild population, and significant genetic differentiation between the wild population and the hatchery samples (F ST=0.058, P < 0.001). These microsatellite loci may be valuable for future population genetic studies, monitoring changes in the genetic variation within stocks in a commercial breeding program, conservation genetics, and molecular assisted selective breeding of the mi-iuy croaker in the future.