Comparative description of ten transcriptomes of newly sequenced invertebrates and efficiency estimation of genomic sampling in non-model taxa

Abstract

IntroductionTraditionally, genomic or transcriptomic data have been restricted to a few model or emerging model organisms, and to a handful of species of medical and/or environmental importance. Next-generation sequencing techniques have the capability of yielding massive amounts of gene sequence data for virtually any species at a modest cost. Here we provide a comparative analysis of de novo assembled transcriptomic data for ten non-model species of previously understudied animal taxa.ResultscDNA libraries of ten species belonging to five animal phyla (2 Annelida [including Sipuncula], 2 Arthropoda, 2 Mollusca, 2 Nemertea, and 2 Porifera) were sequenced in different batches with an Illumina Genome Analyzer II (read length 100 or 150 bp), rendering between ca. 25 and 52 million reads per species. Read thinning, trimming, and de novo assembly were performed under different parameters to optimize output. Between 67,423 and 207,559 contigs were obtained across the ten species, post-optimization. Of those, 9,069 to 25,681 contigs retrieved blast hits against the NCBI non-redundant database, and approximately 50% of these were assigned with Gene Ontology terms, covering all major categories, and with similar percentages in all species. Local blasts against our datasets, using selected genes from major signaling pathways and housekeeping genes, revealed high efficiency in gene recovery compared to available genomes of closely related species. Intriguingly, our transcriptomic datasets detected multiple paralogues in all phyla and in nearly all gene pathways, including housekeeping genes that are traditionally used in phylogenetic applications for their purported single-copy nature.ConclusionsWe generated the first study of comparative transcriptomics across multiple animal phyla (comparing two species per phylum in most cases), established the first Illumina-based transcriptomic datasets for sponge, nemertean, and sipunculan species, and generated a tractable catalogue of annotated genes (or gene fragments) and protein families for ten newly sequenced non-model organisms, some of commercial importance (i.e., Octopus vulgaris). These comprehensive sets of genes can be readily used for phylogenetic analysis, gene expression profiling, developmental analysis, and can also be a powerful resource for gene discovery. The characterization of the transcriptomes of such a diverse array of animal species permitted the comparison of sequencing depth, functional annotation, and efficiency of genomic sampling using the same pipelines, which proved to be similar for all considered species. In addition, the datasets revealed their potential as a resource for paralogue detection, a recurrent concern in various aspects of biological inquiry, including phylogenetics, molecular evolution, development, and cellular biochemistry.

Re-evaluating the phylogeny of Sipuncula through transcriptomics

“…Live specimens of eight species representing all sipunculan families were collected by MJB and GYK: Antillesoma antillarum, Phascolosoma perlucens, Aspidosiphon parvulus, Nephasoma pellucidum, Phascolion cryptum, Siphonosoma cumanenses and Sipunculus nudus (although for this study S. nudus transcriptome was retrieved from Riesgo et al, 2012); Phascolopsis gouldii was obtained from Marine Biological Specimens (Woods Hole, Massachusetts). Information about the sampling localities can be found in the MCZ online collections database (http://mczbase.mcz.harvard.edu) and in Table 1.…”

“…In addition, 10 taxa were chosen as outgroups from which 3 were collected live for this study (Baseodiscus unicolor, Argonomertes australiensis and Chaetopterus sp. ); 3 transcriptomes were retrieved from Riesgo et al (2012; Chiton olivaceus, Hormogaster samnitica and Octopus vulgaris); 2 transcriptomes were retrieved from GenBank (Owenia fusiformis and Magelona johnstoni), and 2 transcriptomes were provided directly by Weigert et al (2014; Eurythoe complanata and Paramphinome jeffreysii).…”

“…For B. unicolor, A. australiensis and Chaetopterus sp., libraries were constructed as described in Riesgo et al (2012). Each library was marked with a distinct index to allow pooling for sequencing.…”

Stress response of the black coral Leiopathes glaberrima when exposed to sub-lethal amounts of crude oil and dispersant

“…The up-regulation of CYP P450 in response to the dispersed oil treatments but not in the bulk oil treatments might indicate that the addition of dispersant increases the availability of oil inside the coral tissue (Zuijdgeest and Huettel, 2012). The fibrillin-2 gene and other genes associated with wound healing (Hemicentins) respond to stress in Nematostella (Reitzel et al, 2008) and other invertebrates (Riesgo et al, 2012). Although experimental cutting of colonies might have also introduced stress (Stossel et al, 2001) and activated genes involved in wound-healing, any general effects of experimental cutting would have been invisible to the differential gene expression analysis because both control and treatment fragments were cut.…”

Construction of PANM Database (Protostome DB) for rapid annotation of NGS data in Mollusks

“…To put it into perspective, the most prolific NGS, the short read length (150 bp) Solexa GAIIx/Illumina platform was able to yield 96,000 Mb reads/run at a reagent cost of $0.12 per Mb (Harrison and Kidner, 2011). With NGS sequencing platforms becoming increasingly cost-efficient with a high-throughput technology model, genetic studies on non-model organisms have improved, enabling the availability of data from the taxons of evolutionary significance (Riesgo et al, 2012).…”