The DNA methylation level is associated with the superior growth of the hybrid crosses in the Pacific oyster Crassostrea gigas

Partial or complete sterility is an obvious feature in triploid Pacific oyster (Crassostrea gigas) which contributes to improving rearing performances. Despite the significance of sterility, the molecular mechanism behind it remains elusive and related research was limited. This study focused on six reproduction-related genes and compared their different behavior in gene expression and DNA methylation pattern between triploid and diploid oysters in order to provide more molecular information. The gonadal development of triploid oyster was examined by histology before molecular analysis. Gametogenesis disturbance was observed in triploid oysters at different development stages (stage II and III) with more serious impairment in females. QPCR showed significant gene expression difference between diploid and triploid in two genes: putative Vg and cgER. Gene expression of putative Vg was delayed in triploids while for cgER triploid oyster showed higher expression and the difference was significant at stage III. DNA methylation pattern of these two genes were further investigated by bisulfite sequencing. Between diploid and triploid oysters, no difference was observed in total methylation level but some individual loci showed different patterns: significantly high methylation rate of loci 2284 in cgER was observed in triploid oyster which has a higher expression of this gene. This study indicated that putative Vg and cgER might play a role in partial sterile in triploid C. gigas. Gene expression could be regulated by the methylation pattern at specific individual locus, which deserves equivalent attention as well as total DNA methylation level.

Salinity is a considerable factor to the survival and distribution for a majority of marine organisms, the drawbacks of which are becoming a serious issue of aquaculture. DNA methylation, an extensively studied epigenetic modification in eukaryotes, plays a significant role in the regulation of gene expression in responding to environmental changes and triggering evolutionary consequences. The Pacific oyster Crassostrea gigas, as a eurythermal and euryhaline species, is considered to be tolerant to salinity fluctuation. In this study, fluorescent-labeled methylation-sensitive amplified polymorphism (F-MSAP) analysis was used to investigate the frequency and variation of DNA methylation in C. gigas under different salinity and time. The results showed that total methylation level was generally on a downward trend. At lower salinity, the total methylation level decreased at the earlier process and then increased during experiment process, but continued to shrink at the rest salinity. Fully methylation tended to better reflect the dynamics of total methylation. Recovery treatment showed that the extent and pattern of DNA methylation were difficult to return to the normal level in this research. The sequencing and BLAST analysis indicated that in salt stress most of the selected bands were closely related to the metabolism of nucleic acids and proteins, tropomyosin, and cellular transport, effecting on different biological processes of C. gigas. This work provides useful data to further elucidate the molecular mechanisms of salt stress response and tolerance in invertebrates.

Effect of inbreeding on performance and genetic parameters of growth and survival traits in the Pacific oyster Crassostrea gigas at larval stage

The basic helix-loop-helix (bHLH) genes encode a large superfamily of transcription factors in the Pacific oyster (Crassostrea gigas), and play a very important role in regulation of growth and development. To investigate the oyster growth traits and the associations with bHLH genes variations, we analyzed the gene polymorphisms-traits association in a wild population, and confirmed the results in another independent wild population by targeted gene re-sequencing and SNPshot analysis. After screening the single nucleotide polymorphisms (SNPs) in three candidate genes of the bHLH family (88 bHLH genes in two wild oyster populations in total), we identified the allele CgLoblHLH4-T/G located in the exon of the CgLoblHLH4 gene. This allele is a non-synonymous mutation (Phe/Leu) with an extremely significant association with shell width (P<0.01) and allele G is beneficial to shell width. This SNP on the CgLoblHLH4 gene might have a potential value as a genetic marker of growth traits that could be used in breeding in C. gigas in the future.

The effect of interstrain hybridization on the production performance in the Pacific oyster Crassostrea gigas

Introduction and evaluation on the US West Coast of a new strain (Midori) of Pacific oyster (Crassostrea gigas) collected from the Ariake Sea, southern Japan

The Pacific oyster Crassostrea gigas has been introduced widely and massively and became an economically important aquaculture species on a global scale. We estimated heritabilities of growth and shell color traits and their genetic correlations in black shell strain of C. gigas. Analyses were performed on 22 full-sib families in a nested mating design including 410 individuals at harvest (24months of age). The parentage assignment was inferred based on four panels of multiplex PCR markers including 10 microsatellite loci and 94.9% of the offspring were unambiguously assigned to single parent pairs. The Spearman correlation test (r=-0.992, P<0.001) demonstrated the high consistency of the shell pigmentation (SP) and L* and their same efficacy in shell color measurements. The narrow-sense heritability estimated under the animal model analysis was 0.18±0.12 for shell height, 0.25±0.16 for shell length, 0.10±0.09 for shell width, 0.42±0.20 for total weight, 0.32±0.18 for shell weight, and 0.68±0.16 for L*, 0.69±0.16 for shell pigmentation, respectively. The considerable additive genetic variation in growth and shell color traits will make it feasible to produce genetic improvements for these traits in selective breeding program. High genetic and phenotypic correlations were found among growth traits and among shell color traits. To optimize a selection strategy for both fast growth and pure dark shell strain of C. gigas, it is proposed to take both total weight and black shell as joint objective traits in selective breeding program. Our study offers an important reference in the process of selective breeding in black shell color stain of C. gigas and will facilitate to develop favorable breeding strategies of genetic improvements for this economically important strain.

Heterosis and genetic diversity of intraspecific hybrids crosses between two selected lines of the Pacific oyster Crassostrea gigas

Genome-wide analysis of the cytochrome P450 gene family in Pacific oyster Crassostrea gigas and their expression profiles during gonad development

First report of a putative involvement of the NMDA pathway in Pacific oyster (Crassostrea gigas) development: Effect of NMDA receptor ligands on oyster metamorphosis with implications for bivalve hatchery management

64. Comparative cryopreservation study of trochophore larvae from two species of bivalves: Pacific oyster (Crassostrea gigas) and blue mussel (Mytilus galloprovinciallis)

Polymorphism in the insulin-related peptide gene and its association with growth traits in the Pacific oyster Crassostrea gigas

BackgroundAlthough bivalves are among the most-studied marine organisms because of their ecological role and economic importance, very little information is available on the genome sequences of oyster species. This report documents three large-scale cDNA sequencing projects for the Pacific oyster Crassostrea gigas initiated to provide a large number of expressed sequence tags that were subsequently compiled in a publicly accessible database. This resource allowed for the identification of a large number of transcripts and provides valuable information for ongoing investigations of tissue-specific and stimulus-dependant gene expression patterns. These data are crucial for constructing comprehensive DNA microarrays, identifying single nucleotide polymorphisms and microsatellites in coding regions, and for identifying genes when the entire genome sequence of C. gigas becomes available.DescriptionIn the present paper, we report the production of 40,845 high-quality ESTs that identify 29,745 unique transcribed sequences consisting of 7,940 contigs and 21,805 singletons. All of these new sequences, together with existing public sequence data, have been compiled into a publicly-available Website http://public-contigbrowser.sigenae.org:9090/Crassostrea_gigas/index.html. Approximately 43% of the unique ESTs had significant matches against the SwissProt database and 27% were annotated using Gene Ontology terms. In addition, we identified a total of 208 in silico microsatellites from the ESTs, with 173 having sufficient flanking sequence for primer design. We also identified a total of 7,530 putative in silico, single-nucleotide polymorphisms using existing and newly-generated EST resources for the Pacific oyster.ConclusionA publicly-available database has been populated with 29,745 unique sequences for the Pacific oyster Crassostrea gigas. The database provides many tools to search cleaned and assembled ESTs. The user may input and submit several filters, such as protein or nucleotide hits, to select and download relevant elements. This database constitutes one of the most developed genomic resources accessible among Lophotrochozoans, an orphan clade of bilateral animals. These data will accelerate the development of both genomics and genetics in a commercially-important species with the highest annual, commercial production of any aquatic organism.

Norgestrel shows androgenic properties and triggers reproductive neuroendocrine toxicity in the testes of Pacific oysters (Crassostrea gigas).

Molecular confirmation and functional study of signal transducer and activator of transcription genes in the Pacific oyster Crassostrea gigas