Background: Comparative genomics approaches can shed light on evolutionary processes that shape differentiation among lineages. The nine-spined stickleback (Pungitius pungitius), a closely related species of the ecological ‘supermodel’ three-spined stickleback (Gasterosteus aculeatus), is an equally promising model system for evolutionary biology research but has as yet garnered less attention and lacks extensive genomic resources. To expand on these resources and improve the study of sticklebacks in a phylogeneticframework, we characterized nine-spined stickleback transcriptomes from brain and liver using massively parallel pyrosequencing.
Results: We obtained nearly eight thousand assembled transcripts, of which 3,091 were assigned as a putative one-to-one ortholog gene found in the three-spined stickleback. These sequences were used for evaluating overall differentiation and substitution rates between nine- and three-spined sticklebacks, and to identify genes that are putatively evolving under positive selection. The synonymous substitution rate was estimated to be 7.1 × 10-9 per site per year between the two species, and a total of 165 genes showed evidence for adaptive evolution in one or both species. A few nine-spined stickleback contigs lacked an obvious ortholog in three-spined sticklebacks but were found to match genes in other fish species, suggesting several gene losses within a few million years since divergence of the two stickleback species. Out of 1,814 SNPs in the nine-spined stickleback contigs (0.044% of evaluated genic sites), we identified 47 SNPs in 25 different genes that differentiate pond and marine ecotypes and found both same and different genes associated with marine and freshwater divergence across sticklebacks. We also identified 468 microsatellites that could be further developed as genetic markers in nine-spined sticklebacks.
Conclusion: With high-throughput pryrosequencing of nine-spined stickleback cDNA libraries, our study provides a significant increase in the number of gene sequences and microsatellite markers for this species, and identifies a number of genes showing evidence of adaptive evolution between nine- and three-spined sticklebacks. We also report several candidate genes that might be involved in differential adaptation between marine and freshwater nine-spined sticklebacks. This study provides a valuable resource for future studies, including the facilitation of studies on this and other stickleback species and identification of candidate genes underlying ecological adaptation of sticklebacks.