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A Genetic linkage map of Atlantic halibut (Hippoglossus hippoglossus L.).

ABSTRACT: A genetic linkage map has been constructed for Atlantic halibut on the basis of 258 microsatellites and 346 AFLPs. Twenty-four linkage groups were identified, consistent with the 24 chromosomes seen in chromosome spreads. The total map distance is 1562.2 cM in the female and 1459.6 cM in the male with an average resolution of 4.3 and 3.5 cM, respectively. Using diploid gynogens, we estimated centromere locations in 19 of 24 linkage groups. Overall recombination in the female was approximately twice that of the male; however, this trend was not consistent along the linkage groups. In the centromeric regions, females had 11-17.5 times the recombination of the males, whereas this trend reversed toward the distal end with males having three times the recombination of the females. Correspondingly, in the male, markers clustered toward the centromeric region with 50% of markers within 20 cM of the putative centromere, whereas 35% of markers in the female were found between 60 and 80 cM from the putative centromere. Limited interspecies comparisons within Japanese flounder and Tetraodon nigroviridis revealed blocks of conservation in sequence and marker order, although regions of chromosomal rearrangement were also apparent.

SUBMITTER: Reid DP

PROVIDER: S-EPMC2034623 | biostudies-literature | 2007 Oct

REPOSITORIES: biostudies-literature

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A Genetic linkage map of Atlantic halibut (Hippoglossus hippoglossus L.).

Reid Darrin P DP Smith Cheryl-Anne CA Rommens Melissa M Blanchard Brian B Martin-Robichaud Debbie D Reith Michael M

Genetics 20070824 2

A genetic linkage map has been constructed for Atlantic halibut on the basis of 258 microsatellites and 346 AFLPs. Twenty-four linkage groups were identified, consistent with the 24 chromosomes seen in chromosome spreads. The total map distance is 1562.2 cM in the female and 1459.6 cM in the male with an average resolution of 4.3 and 3.5 cM, respectively. Using diploid gynogens, we estimated centromere locations in 19 of 24 linkage groups. Overall recombination in the female was approximately tw ...[more]

PMID: 17720928

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Project description:BackgroundFlatfish metamorphosis is a hormone regulated post-embryonic developmental event that transforms a symmetric larva into an asymmetric juvenile. In altricial-gastric teleost fish, differentiation of the stomach takes place after the onset of first feeding, and during metamorphosis dramatic molecular and morphological modifications of the gastrointestinal (GI-) tract occur. Here we present the functional ontogeny of the developing GI-tract from an integrative perspective in the pleuronectiforme Atlantic halibut, and test the hypothesis that the multiple functions of the teleost stomach develop synchronously during metamorphosis.ResultsOnset of gastric function was determined with several approaches (anatomical, biochemical, molecular and in vivo observations). In vivo pH analysis in the GI-tract lumen combined with quantitative PCR (qPCR) of α and β subunits of the gastric proton pump (H+/K+-ATPase) and pepsinogen A2 indicated that gastric proteolytic capacity is established during the climax of metamorphosis. Transcript abundance of ghrelin, a putative orexigenic signalling molecule produced in the developing stomach, correlated (p < 0.05) with the emergence of gastric proteolytic activity, suggesting that the stomach's role in appetite regulation occurs simultaneously with the establishment of proteolytic function. A 3D models series of the GI-tract development indicated a functional pyloric sphincter prior to first feeding. Observations of fed larvae in vivo confirmed that stomach reservoir function was established before metamorphosis, and was thus independent of this event. Mechanical breakdown of food and transportation of chyme through the GI-tract was observed in vivo and resulted from phasic and propagating contractions established well before metamorphosis. The number of contractions in the midgut decreased at metamorphic climax synchronously with establishment of the stomach's proteolytic capacity and its increased peristaltic activity. Putative osmoregulatory competence of the GI-tract, inferred by abundance of Na+/K+-ATPase α transcripts, was already established at the onset of exogenous feeding and was unmodified by metamorphosis.ConclusionsThe functional specialization of the GI-tract was not exclusive to metamorphosis, and its osmoregulatory capacity and reservoir function were established before first feeding. Nonetheless, acid production and the proteolytic capacity of the stomach coincided with metamorphic climax, and also marked the onset of the stomach's involvement in appetite regulation via ghrelin.

Project description:BackgroundAtlantic halibut (Hippoglossus hippoglossus) is a high-value, niche market species for cold-water marine aquaculture. Production of monosex female stocks is desirable in commercial production since females grow faster and mature later than males. Understanding the sex determination mechanism and developing sex-associated markers will shorten the time for the development of monosex female production, thus decreasing the costs of farming.ResultsHalibut juveniles were masculinised with 17 α-methyldihydrotestosterone (MDHT) and grown to maturity. Progeny groups from four treated males were reared and sexed. Two of these groups (n = 26 and 70) consisted of only females, while the other two (n = 30 and 71) contained balanced sex ratios (50% and 48% females respectively). DNA from parents and offspring from the two mixed-sex families were used as a template for Restriction-site Associated DNA (RAD) sequencing. The 648 million raw reads produced 90,105 unique RAD-tags. A linkage map was constructed based on 5703 Single Nucleotide Polymorphism (SNP) markers and 7 microsatellites consisting of 24 linkage groups, which corresponds to the number of chromosome pairs in this species. A major sex determining locus was mapped to linkage group 13 in both families. Assays for 10 SNPs with significant association with phenotypic sex were tested in both population data and in 3 additional families. Using a variety of machine-learning algorithms 97% correct classification could be obtained with the 3% of errors being phenotypic males predicted to be females.ConclusionAltogether our findings support the hypothesis that the Atlantic halibut has an XX/XY sex determination system. Assays are described for sex-associated DNA markers developed from the RAD sequencing analysis to fast track progeny testing and implement monosex female halibut production for an immediate improvement in productivity. These should also help to speed up the inclusion of neomales derived from many families to maintain a larger effective population size and ensure long-term improvement through selective breeding.

Project description:BACKGROUND:The commercial production of Atlantic halibut (Hippoglossus hippoglossus L.) suffers from a major bottleneck due to the low success of producing juveniles for on-growing. Atlantic halibut females are routinely hand-stripped and incorrect timing of stripping can result in low quality eggs due to post-ovulatory aging. Post-ovulatory aging leads to compositional changes in eggs that include maternally provided proteins and RNAs. There have been few studies of the maternally provided mRNA transcripts that control early development in commercially important fish species. The present study aimed to study maternal gene expression in Atlantic halibut and its relation to egg quality parameters including blastomere symmetry and hatching success. RESULTS:A maternal EST library containing 2341 sequences was constructed by suppressive subtractive hybridisation. Thirty genes were selected for expression studies; 23 novel genes and 7 genes with documented roles in early development. The expressions of twenty-one selected genes were measured by qPCR from fertilization to the 10-somite stage. Three genes were identified as strictly maternal genes that were expressed until the start of gastrulation; askopos (kop), si:dkey-30j22.9 (Tudor family member), and Tudor 5 protein (Tdrd5). The expressions of 18 genes at the 8-cell stage were correlated with egg quality parameters. The majority of genes showed either no or very minor correlations with egg quality parameter. However, two genes correlated positively with hatching success (r> 0.50, HHC00353: r = 0.58, p < 0.01; HHC01517: r = 0.56, p < 0.01) and one gene (HHC00255) was negatively correlated with the percentage of normal blastomeres (r = -0.62, p < 0.05). CONCLUSIONS:During this study we have related maternal levels of gene expression to hatching success in fish. Poor hatching success was not correlated with a general decrease in transcript abundance but with low transcript levels of some specific genes. Thus, the molecular mechanisms leading to low Atlantic halibut egg quality cannot be entirely explained by post-ovulatory aging.

Project description:Real time RT-PCR has become an important tool for analyzing gene expression in fish. Although several housekeeping genes have been evaluated in Atlantic halibut (Hippoglossus Hippoglossus L.), appropriate reference genes for low copy mRNA transcripts at the earliest developmental stages have not been identified. No attempts have been reported to identify suitable reference genes in halibut infected with NNV or in stimulated halibut leucocytes. In this study, beta-actin1 (ACTB1), elongation factor 1 alpha (EF1A1), hypoxanthine-guanine phosphoribosyltransferase 1 (HPRT1), ribosomal protein L7 (RPL7), tubulin beta 2C (Tubb2C), and ubiquitin-conjugating enzyme (UbcE) were evaluated as reference genes for normalization of real time RT-PCR data during Atlantic halibut development, in tissue of healthy and NNV-infected fish, and in in vivo and in vitro stimulated anterior kidney leucocytes.The expression of all six genes was relatively stable from the unfertilized egg until 12 day degrees post fertilization (ddpf). However, none of the selected genes were found to be stably expressed throughout halibut development. The mRNA levels of the six genes increased from 18 ddpf, when zygotic transcription is likely to be activated, and stabilized at different time points. The Excel-based software programs BestKeeper, geNorm, and NormFinder ranked EF1A1 and UbcE as the best candidate reference genes before activation of zygotic transcription, and RPL7 and EF1A1 as the best candidates after hatching. EF1A1 and RPL7 were also listed as the best reference genes when exploring the expression levels of the six genes in various halibut organs, both in non-injected fish and in mock- and NNV-injected fish. None of the reference genes were found optimal for normalization of real time RT-PCR data from in vitro stimulated anterior kidney leucocytes.Generally, it was found that EF1A1 and RPL7 were the genes that showed least variation, with HPRT1 and UbcE as intermediate genes, and ACTB1 and Tubb2C as the least stable ones. None of the six reference genes can be recommended as reference gene candidates in ConA-PMA stimulated leucocytes. However, UbcE can be a good candidate in other experimental setups. This study emphasizes the need for reference gene evaluation, as universal reference genes have not been identified.

Project description:BackgroundAtlantic cod (Gadus morhua) is a species with increasing economic significance for the aquaculture industry. The genetic improvement of cod will play a critical role in achieving successful large-scale aquaculture. While many microsatellite markers have been developed in cod, the number of single nucleotide polymorphisms (SNPs) is currently limited. Here we report the identification of SNPs from sequence data generated by a large-scale expressed sequence tag (EST) program, focusing on fish originating from Canadian waters.ResultsA total of 97976 ESTs were assembled to generate 13448 contigs. We detected 4753 SNPs that met our selection criteria (depth of coverage > or = 4 reads; minor allele frequency > 25%). 3072 SNPs were selected for testing. The percentage of successful assays was 75%, with 2291 SNPs amplifying correctly. Of these, 607 (26%) SNPs were monomorphic for all populations tested. In total, 64 (4%) of SNPs are likely to represent duplicated genes or highly similar members of gene families, rather than alternative alleles of the same gene, since they showed a high frequency of heterozygosity. The remaining polymorphic SNPs (1620) were categorised as validated SNPs. The mean minor allele frequency of the validated loci was 0.258 (+/- 0.141). Of the 1514 contigs from which validated SNPs were selected, 31% have a significant blast hit. For the SNPs predicted to occur in coding regions (141), we determined that 36% (51) are non-synonymous. Many loci (1033 SNPs; 64%) are polymorphic in all populations tested. However a small number of SNPs (184) that are polymorphic in the Western Atlantic were monomorphic in fish tested from three European populations. A preliminary linkage map has been constructed with 23 major linkage groups and 924 mapped SNPs.ConclusionsThese SNPs represent powerful tools to accelerate the genetic improvement of cod aquaculture. They have been used to build a genetic linkage map that can be applied to quantitative trait locus (QTL) discovery. Since these SNPs were generated from ESTs, they are linked to specific genes. Genes that map within QTL intervals can be prioritized for testing to determine whether they contribute to observed phenotypes.

Dataset Information

A Genetic linkage map of Atlantic halibut (Hippoglossus hippoglossus L.).

Publications

A Genetic linkage map of Atlantic halibut (Hippoglossus hippoglossus L.).

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