BackgroundBivalves comprise 30,000 extant species, constituting the second largest group of mollusks. However, limited genetic research has focused on this group of animals so far, which is, in part, due to the lack of genomic resources. The advent of high-throughput sequencing technologies enables generation of genomic resources in a short time and at a minimal cost, and therefore provides a turning point for bivalve research. In the present study, we performed de novo transcriptome sequencing to first produce a comprehensive expressed sequence tag (EST) dataset for the Yesso scallop (Patinopecten yessoensis).ResultsIn a single 454 sequencing run, 805,330 reads were produced and then assembled into 32,590 contigs, with about six-fold sequencing coverage. A total of 25,237 unique protein-coding genes were identified from a variety of developmental stages and adult tissues based on sequence similarities with known proteins. As determined by GO annotation and KEGG pathway mapping, functional annotation of the unigenes recovered diverse biological functions and processes. Transcripts putatively involved in growth, reproduction and stress/immune-response were identified. More than 49,000 single nucleotide polymorphisms (SNPs) and 2,700 simple sequence repeats (SSRs) were also detected.ConclusionOur data provide the most comprehensive transcriptomic resource currently available for P. yessoensis. Candidate genes potentially involved in growth, reproduction, and stress/immunity-response were identified, and are worthy of further investigation. A large number of SNPs and SSRs were also identified and ready for marker development. This resource should lay an important foundation for future genetic or genomic studies on this species.

BackgroundSea cucumbers are a special group of marine invertebrates. They occupy a taxonomic position that is believed to be important for understanding the origin and evolution of deuterostomes. Some of them such as Apostichopus japonicus represent commercially important aquaculture species in Asian countries. Many efforts have been devoted to increasing the number of expressed sequence tags (ESTs) for A. japonicus, but a comprehensive characterization of its transcriptome remains lacking. Here, we performed the large-scale transcriptome profiling and characterization by pyrosequencing diverse cDNA libraries from A. japonicus.ResultsIn total, 1,061,078 reads were obtained by 454 sequencing of eight cDNA libraries representing different developmental stages and adult tissues in A. japonicus. These reads were assembled into 29,666 isotigs, which were further clustered into 21,071 isogroups. Nearly 40% of the isogroups showed significant matches to known proteins based on sequence similarity. Gene ontology (GO) and KEGG pathway analyses recovered diverse biological functions and processes. Candidate genes that were potentially involved in aestivation were identified. Transcriptome comparison with the sea urchin Strongylocentrotus purpuratus revealed similar patterns of GO term representation. In addition, 4,882 putative orthologous genes were identified, of which 202 were not present in the non-echinoderm organisms. More than 700 simple sequence repeats (SSRs) and 54,000 single nucleotide polymorphisms (SNPs) were detected in the A. japonicus transcriptome.ConclusionPyrosequencing was proven to be efficient in rapidly identifying a large set of genes for the sea cucumber A. japonicus. Through the large-scale transcriptome sequencing as well as public EST data integration, we performed a comprehensive characterization of the A. japonicus transcriptome and identified candidate aestivation-related genes. A large number of potential genetic markers were also identified from the A. japonicus transcriptome. This transcriptome resource would lay an important foundation for future genetic or genomic studies on this species.

Genetic linkage maps are critical and indispensable tools in a wide range of genetic and genomic research. With the advancement of genotyping-by-sequencing (GBS) methods, the construction of a high-density and high-resolution linkage maps has become achievable in marine organisms lacking sufficient genomic resources, such as echinoderms. In this study, high-density, high-resolution genetic map was constructed for a sea cucumber species, Apostichopus japonicus, utilizing the 2b-restriction site-associated DNA (2b-RAD) method. A total of 7839 markers were anchored to the linkage map with the map coverage of 99.57%, to our knowledge, this is the highest marker density among echinoderm species. QTL mapping and association analysis consistently captured one growth-related QTL located in a 5 cM region of linkage group (LG) 5. An annotated candidate gene, retinoblastoma-binding protein 5 (RbBP5), which has been reported to be an important regulator of cell proliferation, was recognized in the QTL region. This linkage map represents a powerful tool for research involving both fine-scale QTL mapping and marker assisted selection (MAS), and will facilitate chromosome assignment and improve the whole-genome assembly of sea cucumber in the future.

BackgroundBivalves play an important role in the ecosystems they inhabit and represent an important food source all over the world. So far limited genetic research has focused on this group of animals largely due to the lack of sufficient genetic or genomic resources. Here, we performed de novo transcriptome sequencing to produce the most comprehensive expressed sequence tag resource for Zhikong scallop (Chlamys farreri), and conducted the first transcriptome comparison for scallops.ResultsIn a single 454 sequencing run, 1,033,636 reads were produced and then assembled into 26,165 contigs. These contigs were then clustered into 24,437 isotigs and further grouped into 20,056 isogroups. About 47% of the isogroups showed significant matches to known proteins based on sequence similarity. Transcripts putatively involved in growth, reproduction and stress/immune-response were identified through Gene ontology (GO) and KEGG pathway analyses. Transcriptome comparison with Yesso scallop (Patinopecten yessoensis) revealed similar patterns of GO representation. Moreover, 38 putative fast-evolving genes were identified through analyzing the orthologous gene pairs between the two scallop species. More than 46,000 single nucleotide polymorphisms (SNPs) and 350 simple sequence repeats (SSRs) were also detected.ConclusionOur study provides the most comprehensive transcriptomic resource currently available for C. farreri. Based on this resource, we performed the first large-scale transcriptome comparison between the two scallop species, C. farreri and P. yessoensis, and identified a number of putative fast-evolving genes, which may play an important role in scallop speciation and/or local adaptation. A large set of single nucleotide polymorphisms and simple sequence repeats were identified, which are ready for downstream marker development. This transcriptomic resource should lay an important foundation for future genetic or genomic studies on C. farreri.

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Sixteen polymorphic microsatellite markers were developed from a partial genomic library enriched using AG and AC repeats in mud snail (Bullacta exarata). Polymorphism of these loci was screened using 48 adult individuals of B. exarata. The number of alleles per locus ranged from 2 to 13. The observed and expected heterozygosities ranged from 0.5106 to 0.8125 and from 0.4868 to 0.9208, respectively. One locus departed significantly from Hardy-Weinberg equilibrium after Bonferroni correction. No linkage disequilibrium was found between any pair of loci. These polymorphic microsatellite markers are expected to provide valuable genetic information in population structure and genetic diversity, which will certainly facilitate management strategies of the mud snail.Keywords Microsatellite markers Á Bullacta exarata Á Mud snail Á PolymorphismThe mud snail (Bullacta exarata), which is an androgynous molluscan species, is widely distributed along the coasts of West Pacific Ocean in China, Japan and Korea (Ye and Lu 2001). It is considered as an economically important fishery species in eastern China (Ying et al. 2004), and its farming area is expanding continuously year by year. However, the recent monitoring data from the State Oceanic Administration indicates that the mud snail introduced from Jiangsu province has become an invasive species in Laizhou Bay and more than eighty percent of the inter-tidal beach has been occupied. Moreover, it has substituted for native species and become the dominant one along the coast of Laizhou Bay, and the maximum density is more than 160 individuals per square meter. However, the population genetic structure and genetic diversity have not been very well studied to provide basic research background for the management of biological invasions, mainly because of the lack of robust molecular markers. Therefore, we have developed sixteen highly polymorphic microsatellite markers for the mud snail.A genomic library enriched using AG and AC repeats was constructed following the methods described by Karagyozov et al. (1993) and Zhan et al. (2007) with some minor modifications. Briefly, the genomic DNA was extract from one adult mud snail using CTAB-based method (Zhan et al. 2007). The genomic DNA was digested with HaeIII and the fragments with the size of 400-1,200 bp were recovered from a 1.0% low-melting-point agarose gel. A synthesized adaptor (Zhan et al. 2007) were ligated to the size-selected DNA fragments using T4 DNA ligase. Fragments containing microsatellite DNA were captured by hybridization to nylon membranes fixed with oligonucleotides (AG) 15 and (AC) 15 probes. The membrane was washed twice in 29 SSC, 1% SDS for 15 min at 60°C, once in 29 SSC, 1% SDS for 30 min at 65°C and once in 0.59 SSC, 1% SDS for 20 min at 100°C and the other procedures were performed according to the methods described by Zhan et al. (2007). The enriched DNA fragments were ligated into pMD18-T vector (TaKaRa), and the ligation product was transformed into competent Escherichia coli DH5a cells. The recombin...

A total of 5728 sequences from the expressed sequence tag database of sea cucumber Apostichopus japonicus were mined for microsatellite sequences. A total of 151 ESTs containing microsatellites were identified. Of 93 primer pairs designed, 56 amplified expected products and 38 showed polymorphism in 48 individuals tested. The number of alleles ranged from 2 to 10, and the observed and expected heterozygosities varied from 0.000 to 0.900 and from 0.332 to 0.884, respectively. No significant linkage disequilibrium (LD) between pairs of loci was found and 21 loci significantly deviated from the Hardy-Weinberg equilibrium (HWE).