Population genetic analysis and conservation strategies for redtail shrimp Fenneropenaeus penicillatus using ten microsatellite markers.

Fenneropenaeus penicillatus, which is on the Red List of Endangered Species for China, is an important shrimp species. However, there is not enough genetic information on F. penicillatus for conservation and management purposes. Ten microsatellite markers were used to analyze the genetic diversity, genetic differentiation, and population structure of F. penicillatus to provide scientific information for the conservation of the species. Low genetic diversity and moderate genetic differentiation were found among 12 putative populations [Beihai, Dongshan (DS), Hainan (HN), Lianjiang, Nanao (NA), Ningde (ND), Putian, Quanzhou (QZ), Xiamen (XM), Shenzhen, Zhanjiang, and Zhangpu] along the southeast coast of China. QZ, XM, and DS exhibited the highest genetic diversity, while NA and ND had the lowest genetic diversity. Genetic differentiation among all populations, except HN, was low compared to the genetic differentiation between HN and the other 11 putative populations. These 12 putative populations were divided into two subgroups. One group consisted of XM, DS, and QZ. The other group consisted of the other eight putative populations with the exception of HN. The HN Island population requires further study due to its large genetic distance from the other 11 putative populations. Problems with the current conservation strategy are pointed out and suggestions given based on genetic information.

Screening and characterization of novel microsatellite loci in Lateolabrax japonicus.

Lateolabrax japonicus, an economically important species, is widely consumed in the offshore coasts of China, Korea, and Japan. We identified 10 new L. japonicus microsatellite markers, using a modified protocol of fast isolation by AFLP of sequences containing repeats. Thirty L. japonicus individuals were collected from Xiamen, China, to evaluate the degree of polymorphism. The number of identified alleles ranged from three to five. The polymorphism information content varied from 0.267 to 0.711, whereas the observed and expected heterozygosities ranged from 0.249 to 0.706 and 0.294 to 0.751, respectively. One of the 10 loci (L10) deviated from the Hardy-Weinberg equilibrium. These new microsatellite markers will provide a useful tool for the determination of population genetic structure and genetic diversity in L. japonicus.

Development of novel polymorphic microsatellite markers in Siganus fuscescens.

Rabbitfish, Siganus fuscescens, is widely distributed in the Indo-Pacific regions and eastern Mediterranean. Its dwelling place includes reef flats, coral reef regions, and seagrass meadows in tropical area and reef areas or shallow waters in locations at high latitudes. In the present study, 10 new polymorphic microsatellite markers were screened from 30 wild S. fuscescens individuals, using a method of fast isolation protocol and amplified fragment length polymorphism of sequences containing repeats. The number of polymorphic alleles per locus was 3 to 5 with a mean of 4.3, while the value of polymorphic information content ranged from 0.283 to 0.680. The values of the observed and expected heterozygosities were in the range 0.3333-0.8462 and 0.3011-0.7424, respectively. Deviation from Hardy-Weinberg equilibrium was not observed in this study. These polymorphic loci are expected to be effective in evaluating the genetic diversity, population structure, and gene flow and in determining the paternity in S. fuscescens, as well as for conservation management.

Isolation and characterization of microsatellite markers in Ruditapes philippinarum.

Ruditapes philippinarum is considered a commercially valuable species, which is commonly found in tidal flats along West Pacific coasts. In China, it is mainly distributed in the southeast sea. In this study, 16 novel microsatellite loci from the R. philippinarum genome were developed, using the protocol of fast isolation by amplified fragment length polymorphism of sequence containing repeats. Thirty-two wild-caught individuals were used to evaluate the degree of polymorphism of these markers. Our results show that there were 10 polymorphic loci and 6 monomorphic loci. The number of alleles per locus and the polymorphism information content ranged from 2 to 6 and from 0.199 to 0.751, respectively. The observed and expected heterozygosities varied from 0.1333 to 0.6207 and 0.1603 to 0.7412, respectively. Of all loci, only one locus was found to deviate significantly from Hardy-Weinberg equilibrium after Bonferroni correction. The loci identified here will provide useful information for future population genetic studies of R. philippinarum.

Isolation and characterization of eight novel microsatellite markers in Acanthopagrus schlegelii.

Acanthopagrus schlegelii is a warm temperate demersal fish, which inhabits the sediment substrate or rocky reefs in shallow seas. As this fish is a nutritionally endowed species with good palatability, it is a highly valuable commercial species for aquaculture and has a long historical standing in Western Pacific countries. Because the population of this fish is currently declining in China, studies and measures aimed at addressing this decline are needed. In this study, eight microsatellite markers were screened from 30 wild A. schlegelii fishes through the FIASCO method, whereby sequences containing repeats were obtained from amplified fragment length polymorphisms. The allelic number ranged from 3 to 5, with a mean number of 3.625. The average observed heterozygosity was 0.6290, ranging from 0.3214 to 0.8966, while the expected heterozygosity was 0.5435, ranging from 0.3452 to 0.6721. The value for polymorphism information content ranged from 0.313 to 0.666. These results show this population has moderate genetic variation and low genetic diversity. These novel polymorphic loci will be useful for future genetic studies of A. schlegelii.

Isolation and characterization of microsatellite markers in Atrina vexillum Born.

Atrina vexillum Born is an economically valuable species, widely distributed in the coastal waters of temperate and tropical areas of the Asia Pacific region. Twenty one novel microsatellite loci were identified in the genome of A. vexillum Born using the protocol for fast isolation by amplified fragment length polymorphism of sequence containing repeats. Thirty-two wild type individuals were used to evaluate the degree of polymorphism of these markers. We identified 13 polymorphic and 8 monomorphic loci with the number of alleles per locus and the polymorphism information content ranging from 2 to 5 and 0.141 to 0.664, respectively. The observed and expected heterozygosity varied from 0.1250 to 0.7000 and 0.1223 to 0.6216, respectively. Two loci deviated significantly from Hardy-Weinberg equilibrium (HWE) after Bonferroni correction, whereas the other loci were in HWE. These loci are expected to provide useful information for population genetic studies of A. vexillum Born.

Isolation and characterization of novel microsatellite markers in Mercenaria mercenaria.

Mercenaria mercenaria, also known as the hard clam, is widely distributed in the coastal waters of temperate and tropical areas in the Asian Pacific region. This species is widely popular in the international market, especially in the United States, Europe, and other Western countries, because of its high protein value, taste, and simple farming requirements. In this study, 17 novel microsatellite loci from the M. mercenaria genome were developed using the fast isolation by amplified fragment length polymorphism of sequences containing repeats protocol. Thirty-two wild individuals were used to evaluate the degree of polymorphism of these markers. Results indicated that there were 11 polymorphic loci and six monomorphic loci, and the number of alleles per locus and the polymorphism information content ranged from two to six and from 0.059 to 0.498, respectively. The observed and expected heterozygosity varied from 0.0625 to 0.5333 and 0.0615 to 0.4977, respectively. The Y1-4 locus deviated significantly from Hardy-Weinberg equilibrium (HWE) after Bonferroni correction was applied, while the other loci were in HWE. These loci will provide useful information for M. mercenaria population genetic studies.

Isolation and characterization of new microsatellite loci in Fenneropenaeus penicillatus.

Fenneropenaeus penicillatus is one of the major economic shrimp species in China. In this study, 14 novel microsatellite loci were developed using the fast isolation protocol with amplified fragment length polymorphism of sequences containing repeats (FIASCO). Polymorphisms were tested in 30 individuals from a single-wild population. The results showed that the number of alleles at each locus ranged from two to four, and the polymorphism information content varied from 0.314 to 0.692. The observed and expected heterozygosities ranged from 0.3343 to 0.6542 and from 0.3458 to 0.6657, respectively. Three loci deviated significantly from Hardy-Weinberg equilibrium after a Bonferroni correction was applied, while no deviations were detected in the other 11 loci. The new microsatellite loci identified in this study could be useful in future F. penicillatus population genetic, conservation research, population structure assessment, and linkage map construction studies.

Isolation and characterization of new microsatellite markers in red tail prawn, Fenneropenaeus penicillatus, an endangered species in China.

Until recently, Fenneropenaeus penicillatus was considered a commercial shrimp species. However, in 2005, it was included on the Red List as an endangered species by the Chinese government. In this study, 19 new microsatellite markers in F. penicillatus were developed and tested in samples of 32 wild individuals from Nanao, China. Twelve loci were polymorphic and 7 were monomorphic. Of the 12 polymorphic loci, the number of alleles per locus ranged from 3 to 6, with an average of 4.42 alleles per locus. The polymorphism information content ranged from 0.302 to 0.670, with a mean of 0.4817. The observed and expected heterozygosities ranged from 0.2250 to 0.8889 and from 0.1111 to 0.7750, respectively. Significant deviations from Hardy-Weinberg equilibrium (HWE, adjusted P < 0.0042) after a Bonferroni correction were observed in 3 loci (NA-9, NA-57, and NA-64), whereas the other 9 loci were in HWE. These new microsatellite markers will be useful in further research on the population genetic structure of F. penicillatus.

Isolation and characterization of microsatellite loci in hybrid giant tiger grouper.

The hybrid giant tiger grouper is a fish that has considerable commercial value and has become increasingly important for aquaculture in South East Asia since 2008. In order to prevent any reduction in genetic diversity in hybrid grouper as a result of aquaculture, we have identified 21 microsatellite markers that can be used to estimate genetic variation in the fish population. The number of alleles at polymorphic microsatellite loci ranged from 2 to 7, and observed and expected heterozygosities varied from 0.0323 to 0.9643 and 0.0921 to 0.7174, respectively. Polymorphism information content values ranged from 0.088 to 0.737. Nineteen of the 21 loci were in Hardy-Weinberg equilibrium (P > 0.005) after application of the Bonferroni correction (k = 10), the exceptions being ZZLD35 (P < 0.005) and ZZLD36 (P < 0.001). No linkage disequilibrium was detected. These 21 microsatellite markers are potentially of great value for analyzing genetic diversity to provide essential information for sustainable management of these fish.

Identification and characterization of microsatellite markers from the tropical sea cucumber, Stichopus horrens (Selenka).

Tropical commercial sea cucumber Stichopus horrens is extensively distributed throughout the tropical Indo-Pacific region, and wild stocks have been severely depleted over the past decade. In this study, we used the microsatellite enrichment library of S. horrens to identify and characterize 13 microsatellite loci, including 11 polymorphic loci and 2 monomorphic loci. Among the 11 polymorphic loci, the number of alleles was 3-8. The observed and expected heterozygosity varied from 0.1364 to 0.8966 and from 0.1653 to 0.7551, respectively. Additionally, all 11 polymorphic loci showed moderate and high polymorphism with the polymorphism information content (0.271-0.7311). A total of 9 polymorphic loci were in Hardy-Weinberg equilibrium, except for 2 loci (adjusted P = 0.004545). Linkage disequilibrium was not detected in any pairs of polymorphic loci. The present study will be useful for studying genetic structure, population conservation, and breeding of wild S. horrens; moreover, our results contribute to the phylogeny and evolutionary research of Holothuroidea.

Development and characterization of microsatellite markers for molecular genetic diversity in Atrina pectinata.

The pen shell, Atrina pectinata, is an economically valuable species that is widely distributed along the coastal waters of temperate and tropical areas, mainly growing in the Indian and Pacific Oceans. Eight novel microsatellite loci from the genome of A. pectinata were developed using fast isolation by amplified fragment length polymorphism of sequence containing repeats. The loci were screened in 30 wild individuals. The results showed that the number of alleles per locus and the polymorphism information content ranged from 2-6 and from 0.233-0.447, respectively. Observed and expected heterozygosities varied from 0.2069-0.7931 and 0.1887-0.5124, respectively. No significant deviations from Hardy-Weinberg equilibrium were detected. These microsatellite loci will be useful for further population studies of genetic diversity, population structure assessment, and conservation of A. pectinata.

Isolation and characterization of microsatellite markers of sea cucumber Stichopus horrens.

Curry fish (Stichopus horrens) is a tropical holothurian species and is widely distributed in the India-West Pacific. In the present study, 9 polymorphic microsatellite loci were isolated and characterized for S. horrens. These loci were tested in 30 individuals from Hainan Island in China. The number of alleles ranged from 2 to 5. The polymorphism information content ranged from 0.348-0.584. The levels of observed and expected heterozygosities varied from 0.1500-0.8000 and from 0.2014-0.5000, respectively. Most loci were in Hardy-Weinberg equilibrium, except HCS1-27 and HCS2-7, after sequential Bonferroni's correction, and no significant linkage disequilibrium was detected for any pairwise combination of loci. These polymorphic microsatellite loci will be useful for studying population structure and conservation strategy design for S. horrens.

Isolation and characterization of microsatellite loci in Branchiostoma belcheri Gray (Amphioxus).

Branchiostoma belcheri Gray is a second-class, nationally protected protochordate in China. We developed 10 novel polymorphic sites in B. belcheri, which were examined using a population of 30 wild individuals from Xiamen, China. The polymorphism information content ranged between 0.141 and 0.681, and the number of alleles varied from 2 to 5. The expected and observed heterozygosities varied between 0.1528 and 0.6920, and between 0.1429 and 0.5000, respectively. These novel microsatellite markers will facilitate the genetic analysis and protection of wild B. belcheri individuals, and the possible re-stocking of the species in the long-term.

Screening the first set of polymorphic microsatellite loci in Lunella coronata granulata (Turbinidae).

Lunella coronata granulata, from the family Turbini-dae, is an economically important species. The first set of 10 poly-morphic microsatellite loci was screened from L. coronata granulata, and 30 individuals were used to analyze the degree of polymorphism in these loci. The level of observed and expected heterozygosity was 0.0667-0.7333 and 0.0644-0.6628, respectively. The polymorphism information content varied from 0.305 to 0.559. Eight loci were in Hardy-Weinberg equilibrium (HWE) (P > 0.05), while two loci devi-ated significantly from the HWE after Bonferroni's correction (P < 0.005). The isolated microsatellite loci can be utilized in studies of population genetic analysis and they provide important genetic mark-ers for construction of genetic linkage maps and genetic breeding of L. coronata granulata resources.

Isolation and characterization of polymorphic microsatellite loci in the sea cucumber Holothuria scabra.

The sea cucumber Holothuria scabra is an endangered species. In this study, nine new polymorphic microsatellite loci were developed and tested in 30 individuals. The number of alleles ranged from 2 to 5, and the observed and expected heterozygosities ranged from 0.1200 to 0.7391 and from 0.2408 to 0.5983, respectively. No loci significantly deviated from the Hardy-Weinberg equilibrium af-ter a Bonferroni correction, and no significant linkage disequilibrium was found between pairs of loci. These polymorphic microsatellite loci will be useful in studying the genetic diversity of H. scabra and its conservation.

Screening and characterization of novel polymorphic microsatellite markers from sea cucumber Holothuria leucospilota.

The sea cucumber Holothuria leucospilota has high me-dicinal value and rich nutritional edible value, and thus is a commercially important aquatic product in China. Microsatellite loci were developed and screened using a fast isolation protocol and amplified fragment length polymorphism of sequences containing repeats. In this study, 16 novel polymorphic microsatellite markers in H. leucospilota were identified, and the relevant genetic variability index was assessed using 30 individu-als from a wild population. The polymorphic information content ranged from 0.183 to 0.668, and the number of alleles per locus varied from 3 to 5. The observed and expected heterozygosities were 0.0370-0.5000 and 0.0776-0.6250, respectively. With the exception of 3 loci (Y1-15, Y11-1, and Y28), the polymorphic loci were in Hardy-Weinberg equilibrium (P > 0.003125). These polymorphic microsatellite loci will contribute to studies of genetic diversity, the research of population structure, and the design of conservation strategies for H. leucospilota.

Development and characterization of new microsatellite markers of Fenneropenaeus penicillatus.

Thirteen new polymorphic microsatellite markers in Fenneropenaeus penicillatus were isolated and characterized. The polymorphism of the thirteen microsatellite markers was tested by 30 individuals from Lianjiang, China. It showed that the number of al-leles per locus ranged from 3 to 6 and the Polymorphism Information Content (PIC) was from 0.324 to 0.706. The observed and expected heterozygosities were 0.3217-0.8023 and 0.1977-0.6783, respectively. Only one loci (LJ-19) deviated significantly from Hardy-Weinberg equilibrium (HWE) (P < 0.00385) after Bonferroni correction, while the other twelve markers were in HWE after Bonferroni correction (P > 0.00385). The thirteen polymorphic microsatellite markers could pro-vide more genetic data for further research on cultivation and recovery of F. penicillatus.

Isolation and characterization of novel polymorphic microsatellite loci in Atrina vexillum Born (Pinnidae).

The pen shell, Atrina vexillum Born, is an edible shellfish that is widely consumed in the Asia-Pacific region. In this study, 11 polymorphic microsatellite loci were isolated from A. vexillum, and 30 wild individuals were used to evaluate the degree of polymorphism of these markers. The number of alleles per locus ranged from 3 to 8. The polymorphism information content varied from 0.199 to 0.831. The observed and expected heterozygosities were 0.1000-0.8667 and 0.1244-0.8356, respectively. Two loci deviated significantly from the Hardy-Weinberg equilibrium (HWE) after a Bonferroni correction, while the other nine loci were at HWE. These microsatellite loci will be useful in further studies on population genetic analyses, and will provide important genetic data for the conservation and recovery of A. vexillum.

Development and characterization of microsatellite loci in Megalonibea fusca.

Megalonibea fusca is a commercially important large edible fish. In this study, the first set of 10 polymorphic microsatellite loci for M. fusca was developed and characterized. The number of alleles per locus ranged from two to five, with the observed and expected heterozygosities ranging from 0.0667 to 0.7667, and from 0.0644 to 0.5828, respectively. Most of the loci were in Hardy-Weinberg equilibrium (P > 0.05), except for two loci (Mf25 and Mf30) after a Bonferroni's correction (P < 0.005). These informative microsatellite markers will be useful in further studies of the population and conservation genetics of this species.