BackgroundHigh density linkage maps are essential for comparative analysis of synteny, fine mapping of quantitative trait loci (QTL), searching for candidate genes and facilitating genome sequence assembly. However, in most foodfish species, marker density is still low. We previously reported a first generation linkage map with 240 DNA markers and its application to preliminarily map QTL for growth traits in Asian seabass (Lates calcarifer). Here, we report a high-resolution linkage map with 790 microsatellites and SNPs, comparative analysis of synteny, fine-mapping of QTL and the identification of potential candidate genes for growth traits.ResultsA second generation linkage map of Asian seabass was developed with 790 microsatellite and SNP markers. The map spanned a genetic length of 2411.5 cM, with an average intermarker distance of 3.4 cM or 1.1 Mb. This high density map allowed for comparison of the map with Tetraodon nigroviridis genome, which revealed 16 synteny regions between the two species. Moreover, by employing this map we refined QTL to regions of 1.4 and 0.2 cM (or 400 and 50 kb) in linkage groups 2 and 3 in a population containing 380 progeny; potential candidate genes for growth traits in QTL regions were further identified using comparative genome analysis, whose effects on growth traits were investigated. Interestingly, a QTL cluster at Lca371 underlying growth traits of Asian seabass showed similarity to the cathepsin D gene of human, which is related to cancer and Alzheimer's disease.ConclusionsWe constructed a high resolution linkage map, carried out comparative mapping, refined the positions of QTL, identified candidate genes for growth traits and analyzed their effects on growth. Our study developed a framework that will be indispensable for further identification of genes and analysis of molecular variation within the refined QTL to enhance understanding of the molecular basis of growth and speed up genetic improvement of growth performance, and it also provides critical resource for future genome sequence assembly and comparative genomics studies on the evolution of fish genomes.
BackgroundMicroRNAs (miRNAs) play an important role in the regulation of many fundamental biological processes. So far miRNAs have been only identified in a few fish species, although there are over 30,000 fish species living under different environmental conditions on the earth. Here, we described an approach to identify conserved miRNAs and characterized their expression patterns in different tissues for the first time in a food fish species Asian seabass (Lates calcarifer).Methodology/Principal FindingsBy combining a bioinformatics analysis with an approach of homolog-based PCR amplification and sequencing, 63 novel miRNAs belonging to 29 conserved miRNA families were identified. Of which, 59 miRNAs were conserved across 10–86 species (E value≤10−4) and 4 miRNAs were conserved only in fish species. qRT-PCR analysis showed that miR-29, miR-103, miR-125 and several let-7 family members were strongly and ubiquitously expressed in all tissues tested. Interestingly, miR-1, miR-21, miR-183, miR-184 and miR-192 showed highly conserved tissue-specific expression patterns. Exposure of the Asian seabass to lipopolysaccharide (LPS) resulted in up-regulation of over 50% of the identified miRNAs in spleen suggesting the importance of the miRNAs in acute inflammatory immune responses.Conclusions/SignificanceThe approach used in this study is highly effective for identification of conserved miRNAs. The identification of 63 miRNAs and determination of the spatial expression patterns of these miRNAs are valuable resources for further studies on post-transcriptional gene regulation in Asian seabass and other fish species. Further identification of the target genes of these miRNAs would shed new light on their regulatory roles of microRNAs in fish.
Analysis of transcriptomes is of great importance in genomic studies. Asian seabass is an important fish species. A number of genomic tools in it were developed, while large expressed sequence tag (EST) data are lacking. We sequenced ESTs from nine normalized cDNA libraries and obtained 11 431 high-quality ESTs. We retrieved 8524 ESTs from dbEST database and analyzed all 19 975 ESTs using bioinformatics tools. After clustering, we obtained 8837 unique sequences (2838 contigs and 5999 singletons). The average contig length was 574 bp. Annotation of these unique sequences revealed that 48.9% of them showed significant homology to RNA sequences in GenBank. Functional classification of the unique ESTs identified a broad range of genes involved in different functions. We identified 6114 putative single-nucleotide polymorphisms and 634 microsatellites in ESTs. We discovered different temporal and spatial expression patterns of some immune-related genes in the Asian seabass after challenging with a pathogen Vibrio harveyi. The unique EST sequences are being used in developing a cDNA microarray to examine global gene expression and will also facilitate future whole-genome sequence assembly and annotation of Asian seabass and comparative genomics.
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