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. 2016 May 7;17(5):690.
doi: 10.3390/ijms17050690.

Optimizing Hybrid de Novo Transcriptome Assembly and Extending Genomic Resources for Giant Freshwater Prawns (Macrobrachium rosenbergii): The Identification of Genes and Markers Associated with Reproduction

Hyungtaek Jung  1 , Byung-Ha Yoon  2   3 , Woo-Jin Kim  4 , Dong-Wook Kim  5 , David A Hurwood  6 , Russell E Lyons  7 , Krishna R Salin  8 , Heui-Soo Kim  9 , Ilseon Baek  10 , Vincent Chand  11 , Peter B Mather  12
Affiliations

Optimizing Hybrid de Novo Transcriptome Assembly and Extending Genomic Resources for Giant Freshwater Prawns (Macrobrachium rosenbergii): The Identification of Genes and Markers Associated with Reproduction

Hyungtaek Jung et al. Int J Mol Sci. .

Abstract

The giant freshwater prawn, Macrobrachium rosenbergii, a sexually dimorphic decapod crustacean is currently the world's most economically important cultured freshwater crustacean species. Despite its economic importance, there is currently a lack of genomic resources available for this species, and this has limited exploration of the molecular mechanisms that control the M. rosenbergii sex-differentiation system more widely in freshwater prawns. Here, we present the first hybrid transcriptome from M. rosenbergii applying RNA-Seq technologies directed at identifying genes that have potential functional roles in reproductive-related traits. A total of 13,733,210 combined raw reads (1720 Mbp) were obtained from Ion-Torrent PGM and 454 FLX. Bioinformatic analyses based on three state-of-the-art assemblers, the CLC Genomic Workbench, Trans-ABySS, and Trinity, that use single and multiple k-mer methods respectively, were used to analyse the data. The influence of multiple k-mers on assembly performance was assessed to gain insight into transcriptome assembly from short reads. After optimisation, de novo assembly resulted in 44,407 contigs with a mean length of 437 bp, and the assembled transcripts were further functionally annotated to detect single nucleotide polymorphisms and simple sequence repeat motifs. Gene expression analysis was also used to compare expression patterns from ovary and testis tissue libraries to identify genes with potential roles in reproduction and sex differentiation. The large transcript set assembled here represents the most comprehensive set of transcriptomic resources ever developed for reproduction traits in M. rosenbergii, and the large number of genetic markers predicted should constitute an invaluable resource for future genetic research studies on M. rosenbergii and can be applied more widely on other freshwater prawn species in the genus Macrobrachium.

Keywords: Macrobrachium rosenbergii; crustacean; de novo assembly; hybrid transcriptome; optimization; prawn; reproduction.

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Figures

Figure 1
Figure 1
Comparison of de novo assemblies applying multi K-values. K-values in Trinity indicate 25-mer default value. (a) Number of cintigs; (b) Maximum contig length (bp); (c) N50 length (bp), and (d) Average contig length (bp).
Figure 2
Figure 2
Summary of gene ontology (GO) terms for all combined contigs in M. rosenbergii. Similarity of contigs generated with the CLC Genomic Workbench were searched and annotated using WEGO.
Figure 3
Figure 3
Differential gene expression analysis comparing ovary and testis tissues from M. rosenbergii. Red dots indicate significantly differentially expressed genes (transcripts or contigs) and the red curved line is average expression strength in the plot.
See this image and copyright information in PMC

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