Open Access Highly Accessed Research article

Fast skeletal muscle transcriptome of the Gilthead sea bream (Sparus aurata) determined by next generation sequencing

Daniel Garcia de la serrana1*, Alicia Estévez2, Karl Andree2 and Ian A Johnston1*

Author Affiliations

1 Physiological and Evolutionary Genomics Laboratory, Scottish Oceans Institute, School of Biology, University of St Andrews, Fife, KY16 8LB, , Scotland, UK

2 Institute for Aquaculture and Food Technology Research (IRTA), Sant Carles de la Ràpita, P.O. Box 200, 43540, Spain

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BMC Genomics 2012, 13:181  doi:10.1186/1471-2164-13-181

Published: 11 May 2012

Additional files

Additional file 1:

Isotig nucleotide sequences from the gilthead sea bream fast muscle transcriptome. 454 reads were assembled using Newbler version 2.5 after trimming adaptors used for dsDNA synthesis and in silico normalization.

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Additional file 2:

Gilthead sea bream transcriptome sequences annotated. Annotation was performed by Blast2GO software. Sequences were blasted with Blastx algorithms against the NCBI non-redundant protein collection (nr) database with a threshold of 10−3. Annotation was done with an E-value Hit Filter of 10−6 combined with an Annotation Cutoff of 55 and GO weighting of 5.

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Additional file 3:

Gilthead sea bream fast muscle transcriptome annotation results. All graphs and figures were prepared using Blast2GO software (A) Data distribution of the gilthead sea bream transcriptome annotation results. NoBlastHits represents transcripts with no blastx results. NoMapping category represents blasted sequences with no gene ontology (GO) annotation. NoAnnot sequences are isotigs with a preliminary GO annotation (mapping) which failed to arrive to the minimal annotation threefold. Annot barr represents isotigs that were successfully blasted and annotated (B) Annotated transcripts length (bp) distribution (C) Transcripts BLAST results e-value distribution (e-value threefold 10−3). Sequences were blasted by blastx algorithm against the nr protein database from NCBI (D) Species distribution of top hits from whole fast muscle 454 transcriptome with significant homology (<10−3) to searches from the NCBI nr database. Only the best/first sequence alignments for a given Blast result for all blast results are show (E) Percentage of sequences annotated as a function of their length (bp) (F) Distribution of transcripts from gilthead sea bream transcriptome with major categories of level 3 molecular function from GO analysis (G) Distribution of transcripts from gilthead sea bream transcriptome with major categories of level 3 biological process from GO analysis (H) Distribution of transcripts from gilthead sea bream transcriptome with major categories of level 6 cellular component from GO analysis.

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Additional file 4:

Transcripts from fast muscle gilthead sea bream (Sparus aurata L.) 454 transcriptome summarized by their gene ontology annotation (GO) according to Biological Process, Molecular Function and Cellular Component. Table only shows the most abundant GO terms from each category as a percentage and the number of transcripts associated to this level.

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Additional file 5:

Kyoto Encyclopaedia of Genes and Genomes (KEGG) maps present in the gilthead sea bream fast muscle transcriptome. Transcripts were annotated to KEGG maps using the automatic annotator tool KAAS [60].

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Additional file 6:

KEGG annotation results. File contains the KEGG maps where isotigs were successfully mapped. Pathway element in green boxes indicates the components represented by at least, one isotig. Gilthead sea bream isotigs were automatically annotated to KEGG maps using KAAS website [60]. The SBH method, optimized for ESTs annotation, was used against human, chimpanzee, orang-utan, rhesus, mouse, rat, dog, giant panda, cow, pig, horse, opossum, platypus, chicken, clawed frog, zebrafish, fruit fly and nematode pathway databases.

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Additional file 7:

Hierarchical information of the isotigs mapped to the KEGG pathways maps using the automatic annotation tool KAAS. File can be opened using KegHier from [60] when maps are displayed the pathway components with an isotig mapped are labeled in red.

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Additional file 8:

PI3K/Akt and mTOR genes represented in the transcriptome mapped onto a reconstruction of the pathway based on KEGG maps (KO:04150 and KO:04910). AMPKα/β/γ, 5′-AMP-activated protein kinase catalytic subunit alpha/beta/gamma; AKT2, Rac-beta serine/threonine protein kinase 2; AKT3, Rac-beta serine/threonine protein kinase 3; ATG1, Serine/threonine-protein kinase ULK1; CBL-B, E3-Ubiquitin-protein kinase CBL; CIP4 (TRIP10), Thyroid hormone receptor interactor 10; eIF4B, Eukaryotic translation initiation factor 4B; eIF4E, Eukaryotic translation initiation factor 4E; Exo-70, Exocyst complex component 7; GYS, Glycogen synthase; GSK3a, Glycogen synthase kinase 3-alpha; GLUT4, Solute carrier family 2, facilitated glucose transporter member 4; HIF1α, Hypoxia inducible factor 1-alpha; HIF3α, Hypoxia inducible factor 3-alpha; IGF, insulin-like growth factor; IGFBP, insulin-like growth factor binding protein; IRS, insulin receptor substrate; mTOR, Mammalian target of rapamycin; p110, Phosphatidylinositol 3-kinase catalytic subunit; p85, Phosphatidylinositol 3-kinase regulatory subunit; PDPK1, 3-phosphoinositide dependent protein kinase 1; PPP1R, Protein phosphatase 1 regulatory subunit; PPP1C, Protein phosphatase 1 catalytic subunit; PHKG, Phosphorylase b kinase gamma; PHK1-B, Phosphorylase kinase alpha-beta subunit; PPAR, Peroxisome proliferation-activated receptor; PYG, Starch phosphorylase; RICTOR, Rapamycin-insensitive companion of mTOR; RAPTOR, Regulatory-associated protein of mTOR; RPS70K, p70 Ribosomal protein S6 kinase; RHEB2, Ras homolog enriched brain; STRADα, STE20 related kinase adaptor alpha; S6, Ribosomal protein S6; TSC, Tuberous sclerosis; TC10 (RHOQ), Ras homolog gene family member Q; VEGF-A1, Vascular endothelial growth factor A1; v-CRK, Proto-oncogen C-crk; 4EBP1, Eukaryotic translator factor 4E binding protein. Numbers associated with the gene name represents isotig length (bp), isotig mean coverage and percentage of identity with the zebrafish orthologue.

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Additional file 9:

Isotig amino acid sequences from gilthead sea bream transcriptome that covers over 90% of the gene coding sequence. Isoigs were translated to peptides using the Blast2GO ORF translation tool. The percentage of CDS coverage was estimated by blasting translated sequences against the NCBI non-redundant protein database.

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Additional file 10:

Predicted amino acids sequences of splice variants.

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Additional file 11:

Gilthead sea bream fast muscle transcriptome microsatellite summary. Microsatellites were identified using msatcommander-1.0.3-alpha and verified their UTR localization by blastx comparison against the NCBI non-redundant protein (nr) database.

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Additional file 12:

Gilthead sea bream paralogues sequences and their alignments. Alignments have been done using ClustalW.

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Additional file 13:

Phylogenetic analysis of paralogues. The amino acids sequences were blasted against the zebrafish (Danio rerio), stickleback (Gasterosteus aculeatus), Takifugu (Takifugu rubripes), medaka (Oryzias latipes), green puffer fish (Tetraodon nigroviridis), chicken (Gallus gallus), frog (Xenopus laevis) and human (Homo sapiens) genomes using Essembl [63]. Alignment of the sequences was performed using the GUIDANCE web tool [67]. The best evolutionary model was estimated for each alignment using MEGA5 software. Maximum Likelihood phylogenetic analysis was constructed, with the best evolutionary model, using PhylM [69].

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Additional file 14:

Detailed list of transcription related isotigs found in the gilthead sea bream transcriptome.

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Additional file 15:

The 20 most abundant isotigs from gilthead sea bream partial assemblies. Abundant isotigs were defined as the ones with the highest number of reads. Name in red are genes found in all conditions top 20. Name in blue are genes found in only one condition.

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Additional file 16:

Adapters and abundant genes sequences used for assembly trimming and normalisation respectively.

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