Targeted SNP discovery in Atlantic salmon (Salmo salar) genes using a 3'UTR-primed SNP detection approach.

BACKGROUND: Single nucleotide polymorphisms (SNPs) represent the most widespread type of DNA variation in vertebrates and may be used as genetic markers for a range of applications. This has led to an increased interest in identification of SNP markers in non-model species and farmed animals. The in silico SNP mining method used for discovery of most known SNPs in Atlantic salmon (Salmo salar) has applied a global (genome-wide) approach. In this study we present a targeted 3'UTR-primed SNP discovery strategy that utilizes sequence data from Salmo salar full length sequenced cDNAs (FLIcs). We compare the efficiency of this new strategy to the in silico SNP mining method when using both methods for targeted SNP discovery. RESULTS: The SNP discovery efficiency of the two methods was tested in a set of FLIc target genes. The 3'UTR-primed SNP discovery method detected novel SNPs in 35% of the target genes while the in silico SNP mining method detected novel SNPs in 15% of the target genes. Furthermore, the 3'UTR-primed SNP discovery strategy was the less labor intensive one and revealed a higher success rate than the in silico SNP mining method in the initial amplification step. When testing the methods we discovered 112 novel bi-allelic polymorphisms (type I markers) in 88 salmon genes [dbSNP: ss179319972-179320081, ss250608647-250608648], and three of the SNPs discovered were missense substitutions. CONCLUSIONS: Full length insert cDNAs (FLIcs) are important genomic resources that have been developed in many farmed animals. The 3'UTR-primed SNP discovery strategy successfully utilized FLIc data to detect novel SNPs in the partially tetraploid Atlantic salmon. This strategy may therefore be useful for targeted SNP discovery in several species, and particularly useful in species that, like salmonids, have duplicated genomes.

Characterization of full-length sequenced cDNA inserts (FLIcs) from Atlantic salmon (Salmo salar).

BACKGROUND: Sequencing of the Atlantic salmon genome is now being planned by an international research consortium. Full-length sequenced inserts from cDNAs (FLIcs) are an important tool for correct annotation and clustering of the genomic sequence in any species. The large amount of highly similar duplicate sequences caused by the relatively recent genome duplication in the salmonid ancestor represents a particular challenge for the genome project. FLIcs will therefore be an extremely useful resource for the Atlantic salmon sequencing project. In addition to be helpful in order to distinguish between duplicate genome regions and in determining correct gene structures, FLIcs are an important resource for functional genomic studies and for investigation of regulatory elements controlling gene expression. In contrast to the large number of ESTs available, including the ESTs from 23 developmental and tissue specific cDNA libraries contributed by the Salmon Genome Project (SGP), the number of sequences where the full-length of the cDNA insert has been determined has been small. RESULTS: High quality full-length insert sequences from 560 pre-smolt white muscle tissue specific cDNAs were generated, accession numbers [GenBank: BT043497 - BT044056]. Five hundred and ten (91%) of the transcripts were annotated using Gene Ontology (GO) terms and 440 of the FLIcs are likely to contain a complete coding sequence (cCDS). The sequence information was used to identify putative paralogs, characterize salmon Kozak motifs, polyadenylation signal variation and to identify motifs likely to be involved in the regulation of particular genes. Finally, conserved 7-mers in the 3'UTRs were identified, of which some were identical to miRNA target sequences. CONCLUSION: This paper describes the first Atlantic salmon FLIcs from a tissue and developmental stage specific cDNA library. We have demonstrated that many FLIcs contained a complete coding sequence (cCDS). This suggests that the remaining cDNA libraries generated by SGP represent a valuable cCDS FLIc source. The conservation of 7-mers in 3'UTRs indicates that these motifs are functionally important. Identity between some of these 7-mers and miRNA target sequences suggests that they are miRNA targets in Salmo salar transcripts as well.