De novo assembly, characterization and functional annotation of Senegalese sole (Solea senegalensis) and common sole (Solea solea) transcriptomes: integration in a database and design of a microarray

Abstract

BackgroundSenegalese sole (Solea senegalensis) and common sole (S. solea) are two economically and evolutionary important flatfish species both in fisheries and aquaculture. Although some genomic resources and tools were recently described in these species, further sequencing efforts are required to establish a complete transcriptome, and to identify new molecular markers. Moreover, the comparative analysis of transcriptomes will be useful to understand flatfish evolution.ResultsA comprehensive characterization of the transcriptome for each species was carried out using a large set of Illumina data (more than 1,800 millions reads for each sole species) and 454 reads (more than 5 millions reads only in S. senegalensis), providing coverages ranging from 1,384x to 2,543x. After a de novo assembly, 45,063 and 38,402 different transcripts were obtained, comprising 18,738 and 22,683 full-length cDNAs in S. senegalensis and S. solea, respectively. A reference transcriptome with the longest unique transcripts and putative non-redundant new transcripts was established for each species. A subset of 11,953 reference transcripts was qualified as highly reliable orthologs (>97% identity) between both species. A small subset of putative species-specific, lineage-specific and flatfish-specific transcripts were also identified. Furthermore, transcriptome data permitted the identification of single nucleotide polymorphisms and simple-sequence repeats confirmed by FISH to be used in further genetic and expression studies. Moreover, evidences on the retention of crystallins crybb1, crybb1-like and crybb3 in the two species of soles are also presented. Transcriptome information was applied to the design of a microarray tool in S. senegalensis that was successfully tested and validated by qPCR. Finally, transcriptomic data were hosted and structured at SoleaDB.ConclusionsTranscriptomes and molecular markers identified in this study represent a valuable source for future genomic studies in these economically important species. Orthology analysis provided new clues regarding sole genome evolution indicating a divergent evolution of crystallins in flatfish. The design of a microarray and establishment of a reference transcriptome will be useful for large-scale gene expression studies. Moreover, the integration of transcriptomic data in the SoleaDB will facilitate the management of genomic information in these important species.Electronic supplementary materialThe online version of this article (doi:10.1186/1471-2164-15-952) contains supplementary material, which is available to authorized users.

Characterization of Iodine-Related Molecular Processes in the Marine Microalga Tisochrysis lutea (Haptophyta)

“…In the case of microalgae cultivated with chloramphenicol, total RNA from the three biological replicates was also pooled and the cDNA was not normalized but the most abundant bands visible by electrophoreses corresponding to chloroplast sequences were cut and removed before 454 libraries preparation. These samples were sequenced by 454/Roche technology as previously described (Benzekri et al, 2014).…”

“…Secondly, seven samples of the experiment described above to evaluate the effect of H 2 O 2 and iodide on gene expression were selected: untreated control at 5 and 45 min, I − -added (0.5 mM) samples at 5 and 45 min, I − -and H 2 O 2 -added (200 µM) samples at 5 and 45 min and sample H 2 O 2 -added at 5 min. The three biological replicates of each condition (∼2 × 10 8 cells at 4 day of culture) were equally pooled and seven cDNA libraries were prepared using the mRNA-Seq sample preparation kit as previously described (Benzekri et al, 2014).…”

Reproduction, olfaction and dominance behaviour in Senegalese sole (Solea senegalensis)

“…For RNA-seq analysis, Illumina short-reads were pre-processed using SeqTrimNext pipeline (Falgueras et al, 2010) available at the Andalusian Bioinformatics Platform (University of Málaga, Spain) using the specific configuration parameters for illumina data. Clean reads were mapped onto the reference and full transcriptomes v4.1 of S. senegalensis available at SoleaDB (Benzekri et al, 2014) using Bowtie2 (Langmead and Salzberg, 2012). These transcriptomes offer complementary information during statistical analysis since the reference transcriptome has a low level of transcript redundancy whereas the full transcriptome possesses the highest level of transcript representation.…”

“…Only significant data after Benjamin-Hochberg method for multiple testing correction with a P-value cut-off 0.05 were considered. Enrichment Functional analysis was carried using the ClueGO Cytoscape plug-in (Bindea et al, 2009) and the orthology annotations available in the SoleaDB (Benzekri et al, 2014). RNA-seq data has been deposited in the Sequence Read Archive (SRA) database with bioproject number PRJNA319182.…”

“…To assess differential gene expression, pre-processed Illumina short-reads were firstly mapped onto the S. senegalensis reference and full transcriptomes v4.1 available at SoleaDB (Benzekri et al, 2014) and DET were identified by using three distinct statistical methods (DESeq implemented in Robina, TCC and NOISeq). Percentage of reads mapped onto the reference transcriptome ranged between 72.6 and 76.5% and between 91.0-93.5% for full transcriptome.…”