Sequencing and Assembly of the Asian seabass Transcriptome

The Asian seabass (or barramundi; Lates calcarifer) is a carnivorous marine teleost and a popular food fish in Southeast Asia. It has an estimated genome size of ~700Mb which has not been sequenced yet, while its transcriptome is poorly represented by just over 22,000 EST sequences in GenBank. Here, we report the sequencing and assembly of the Asian seabass transcriptome, which will be an essential foundation for the development of tools for gene expression analysis and genome annotation. We have utilized a hybrid approach by sequencing samples pooled from various organs of multiple individuals using three different platforms, namely SOLiD 3+, 454 FLX Titanium and paired-end Illumina HiSeq 2000. The de novo assembly was performed using a combination of Trinity, CLCBio and CAP3. A total of 362,369 transcript sequences, inclusive of alternative splice variants and paralogues were generated, and the first application of this dataset was to design an Asian seabass expression microarray (Agilent 8X60k). Sequences for the microarray design were selected using two approaches, first by using BLASTN/BLASTX to select a representative seabass sequence for every gene in a reference sequence database, and secondly, by mapping the SOLID 3+ male testis and female ovary reads against those transcripts without a BLASTN/BLASTX hit to select sequences that were differentially expressed between the two gonads. This resulted in 61,702 sequences for the microarray, which will be used in our future projects relating to nutrigenomics and disease resistance, as well as analyses of the maturation and transformation of the seabass gonads.