Gene discovery in the Antarctic fur seal (Arctocephalus gazella) skin transcriptome

Abstract

Next-generation sequencing provides a powerful new approach for developing functional genomic tools for nonmodel species, helping to narrow the gap between studies of model organisms and those of natural populations. Consequently, massively parallel 454 sequencing was used to characterize a normalized cDNA library derived from skin biopsy samples of twelve Antarctic fur seal (Arctocephalus gazella) individuals. Over 412 Mb of sequence data were generated, comprising 1.4 million reads of average length 286 bp. De novo assembly using Newbler 2.3 yielded 156 contigs plus 22 869 isotigs, which in turn clustered into 18,576 isogroups. Almost half of the assembled transcript sequences showed significant similarity to the nr database, revealing a functionally diverse array of genes. Moreover, 97.9% of these mapped to the dog (Canis lupis familiaris) genome, with a strong positive relationship between the number of sequences locating to a given chromosome and the length of that chromosome in the dog indicating a broad genomic distribution. Average depth of coverage was also almost 20-fold, sufficient to detect several thousand putative microsatellite loci and single nucleotide polymorphisms. This study constitutes an important step towards developing genomic resources with which to address consequential questions in pinniped ecology and evolution. It also supports an earlier but smaller study showing that skin tissue can be a rich source of expressed genes, with important implications for studying the genomics not only of marine mammals, but also more generally of species that cannot be destructively sampled.

Expression Profiling without Genome Sequence Information in a Non-Model Species, Pandalid Shrimp (Pandalus latirostris), by Next-Generation Sequencing

“…Furthermore, species differentiation of crow species Corvus corone and Corvus cornix was detected through differences in their gene expression profiles, whereas the use of several DNA markers failed to detect this differentiation [18]. In mammals, transcriptome analyses in the Antarctic fur seal Arctocephalus gazella using mRNA derived from skin tissues detected several thousand putative microsatellite loci and SNPs [19]. These pioneering studies have suggested that NGS provides a massive amount of useful information for population genetics in non-model organisms.…”

A Novel Approach for Mining Polymorphic Microsatellite Markers In Silico

“…Just such an opportunity is provided by a large body of 454 sequence data recently generated for the Antarctic fur seal, Arctocephalus gazella [15]. A long-term genetic study of this colonially breeding polygynous pinniped species at Bird Island, South Georgia [16] has identified remarkably consistent relationships between heterozygosity measured at 9–76 microsatellite loci and a variety of important fitness traits, from male reproductive success through body size to attractiveness [17]–[20].…”

Transcriptomic SNP discovery for custom genotyping arrays: impacts of sequence data, SNP calling method and genotyping technology on the probability of validation success

“…This was constructed using 454 sequence reads generated from two different cDNA libraries, one comprising skin samples from 12 individuals [33] and the other comprising necropsy samples from nine individuals [34]. Assembly of these data using NEWBLER generated a total of 23,096 contigs [34], which we refer to as the ‘454 transcriptome’.…”

“…In order to determine the extent of overlap between SNPs called by different methods, we revisited two sets of SNPs called from the 454 transcriptome using NEWBLER and SWAP454 respectively [33, 34]. A small number of SNPs within these datasets were duplicated or had an alternative allele frequency of one.…”

“…A transcriptome assembly based on Roche 454 sequencing is already available for this species, from which two SNP datasets were generated using NEWBLER and SWAP454 respectively [33, 34]. Here, we supplement this transcriptome with short read Illumina sequencing, allowing a comparison of SNP discovery methods tailored to different types of sequence data.…”