Abstract
BackgroundTurbot (Scophthalmus maximus L.) is an important aquacultural resource both in Europe and Asia. However, there is little information on gene sequences available in public databases. Currently, one of the main problems affecting the culture of this flatfish is mortality due to several pathogens, especially viral diseases which are not treatable. In order to identify new genes involved in immune defense, we conducted 454-pyrosequencing of the turbot transcriptome after different immune stimulations.Methodology/Principal FindingsTurbot were injected with viral stimuli to increase the expression level of immune-related genes. High-throughput deep sequencing using 454-pyrosequencing technology yielded 915,256 high-quality reads. These sequences were assembled into 55,404 contigs that were subjected to annotation steps. Intriguingly, 55.16% of the deduced protein was not significantly similar to any sequences in the databases used for the annotation and only 0.85% of the BLASTx top-hits matched S. maximus protein sequences. This relatively low level of annotation is possibly due to the limited information for this specie and other flatfish in the database. These results suggest the identification of a large number of new genes in turbot and in fish in general. A more detailed analysis showed the presence of putative members of several innate and specific immune pathways.Conclusions/SignificanceTo our knowledge, this study is the first transcriptome analysis using 454-pyrosequencing for turbot. Previously, there were only 12,471 EST and less of 1,500 nucleotide sequences for S. maximus in NCBI database. Our results provide a rich source of data (55,404 contigs and 181,845 singletons) for discovering and identifying new genes, which will serve as a basis for microarray construction, gene expression characterization and for identification of genetic markers to be used in several applications. Immune stimulation in turbot was very effective, obtaining an enormous variety of sequences belonging to genes involved in the defense mechanisms.
Highlights
Turbot (Scophthalmus maximus) is an economically important flatfish species belonging to the family Scophthalmidae widely distributed from Norway to the Mediterranean and the Black Sea [1]
Our results provide a rich source of data (55,404 contigs and 181,845 singletons) for discovering and identifying new genes, which will serve as a basis for microarray construction, gene expression characterization and for identification of genetic markers to be used in several applications
Among the colony stimulating factors (CSFs), which regulate the production and functional activities of hematopoietic cells [70], we have found four types: macrophage colonystimulating factor 1 (MCSF-1) and 2 (MCSF-2), granulocyte colony-stimulating factor 1 (GCSF-1) and the megakaryocyte colony-stimulating factor thrombopoietin
Summary
Turbot (Scophthalmus maximus) is an economically important flatfish species belonging to the family Scophthalmidae (order Pleuronectiformes) widely distributed from Norway to the Mediterranean and the Black Sea [1]. In Europe, turbot aquaculture production was 9,067 tons in 2008, representing almost 90% of total flatfish production, being Spain the main European producer [2]. This species, native to Europe, is cultured in China, reaching an annual level of 50,000–60,000 tons in recent years and being the largest producer of turbot in the world [3]. This development caused a parallel increase in pathological conditions affecting this culture. In order to identify new genes involved in immune defense, we conducted 454-pyrosequencing of the turbot transcriptome after different immune stimulations
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