Dataset Information

Comparative Analysis of Strategies for De Novo Transcriptome Assembly in Prokaryotes: Streptomyces clavuligerus as a Case Study.

ABSTRACT: The performance of software tools for de novo transcriptome assembly greatly depends on the selection of software parameters. Up to now, the development of de novo transcriptome assembly for prokaryotes has not been as remarkable as that for eukaryotes. In this contribution, Rockhopper2 was used to perform a comparative transcriptome analysis of Streptomyces clavuligerus exposed to diverse environmental conditions. The study focused on assessing the incidence of software parameters on software performance for the identification of differentially expressed genes as a final goal. For this, a statistical optimization was performed using the Transrate Assembly Score (TAS). TAS was also used for evaluating the software performance and for comparing it with related tools, e.g., Trinity. Transcriptome redundancy and completeness were also considered for this analysis. Rockhopper2 and Trinity reached a TAS value of 0.55092 and 0.58337, respectively. Trinity assembles transcriptomes with high redundancy, with 55.6% of transcripts having some duplicates. Additionally, we observed that the total number of differentially expressed genes (DEG) and their annotation greatly depends on the method used for removing redundancy and the tools used for transcript quantification. To our knowledge, this is the first work aimed at assessing de novo assembly software for prokaryotic organisms.

SUBMITTER: Caicedo-Montoya C

PROVIDER: S-EPMC6970227 | biostudies-literature | 2019 Nov

REPOSITORIES: biostudies-literature

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Comparative Analysis of Strategies for De Novo Transcriptome Assembly in Prokaryotes: Streptomyces clavuligerus as a Case Study.

Caicedo-Montoya Carlos C Pinilla Laura L Toro León F LF Yepes-García Jeferyd J Ríos-Estepa Rigoberto R

High-throughput 20191130 4

The performance of software tools for de novo transcriptome assembly greatly depends on the selection of software parameters. Up to now, the development of de novo transcriptome assembly for prokaryotes has not been as remarkable as that for eukaryotes. In this contribution, Rockhopper2 was used to perform a comparative transcriptome analysis of Streptomyces clavuligerus exposed to diverse environmental conditions. The study focused on assessing the incidence of software parameters on sof ...[more]

PMID: 31801255

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Project description:BACKGROUND: Common carp (Cyprinus carpio) is one of the most important aquaculture species of Cyprinidae with an annual global production of 3.4 million tons, accounting for nearly 14% of the freshwater aquaculture production in the world. Due to the economical and ecological importance of common carp, genomic data are eagerly needed for genetic improvement purpose. However, there is still no sufficient transcriptome data available. The objective of the project is to sequence transcriptome deeply and provide well-assembled transcriptome sequences to common carp research community. RESULT: Transcriptome sequencing of common carp was performed using Roche 454 platform. A total of 1,418,591 clean ESTs were collected and assembled into 36,811 cDNA contigs, with average length of 888 bp and N50 length of 1,002 bp. Annotation was performed and a total of 19,165 unique proteins were identified from assembled contigs. Gene ontology and KEGG analysis were performed and classified all contigs into functional categories for understanding gene functions and regulation pathways. Open Reading Frames (ORFs) were detected from 29,869 (81.1%) contigs with an average ORF length of 763 bp. From these contigs, 9,625 full-length cDNAs were identified with sequence length from 201 bp to 9,956 bp. Comparative analysis revealed that 27,693(75.2%) contigs have significant similarity to zebrafish Refseq proteins, and 24,371(66.2%), 24,501(66.5%) and 25,025(70.0%) to teraodon, medaka and three-spined stickleback refseq proteins. A total of 2,064 microsatellites were initially identified from 1,730 contigs, and 1,639 unique sequences had sufficient flanking sequences on both sides for primer design. CONCLUSION: The transcriptome of common carp had been deep sequenced, de novo assembled and characterized, providing the valuable resource for better understanding of common carp genome. The transcriptome data will facilitate future functional studies on common carp genome, and gradually apply in breeding programs of common carp, as well as closely related other Cyprinids.

Project description:BackgroundThe phytoflagellated protozoan, Euglena gracilis, has been proposed as an attractive feedstock for the accumulation of valuable compounds such as β-1,3-glucan, also known as paramylon, and wax esters. The production of wax esters proceeds under anaerobic conditions, designated as wax ester fermentation. In spite of the importance and usefulness of Euglena, the genome and transcriptome data are currently unavailable, though another research group has recently published E.gracilis transcriptome study during our submission. We herein performed an RNA-Seq analysis to provide a comprehensive sequence resource and some insights into the regulation of genes including wax ester metabolism by comparative transcriptome analysis of E.gracilis under aerobic and anaerobic conditions.ResultsThe E.gracilis transcriptome analysis was performed using the Illumina platform and yielded 90.3 million reads after the filtering steps. A total of 49,826 components were assembled and identified as a reference sequence of E.gracilis, of which 26,479 sequences were considered to be potentially expressed (having FPKM value of greater than 1). Approximately half of all components were estimated to be regulated in a trans-splicing manner, with the addition of protruding spliced leader sequences. Nearly 40 % of 26,479 sequences were annotated by similarity to Swiss-Prot database using the BLASTX program. A total of 2080 transcripts were identified as differentially expressed genes (DEGs) in response to anaerobic treatment for 24 h. A comprehensive pathway enrichment analysis using the KEGG pathway revealed that the majority of DEGs were involved in photosynthesis, nucleotide metabolism, oxidative phosphorylation, fatty acid metabolism. We successfully identified a candidate gene set of paramylon and wax esters, including novel β-1,3-glucan and wax ester synthases. A comparative expression analysis of aerobic- and anaerobic-treated E.gracilis cells indicated that gene expression changes in these components were not extensive or dynamic during the anaerobic treatment.ConclusionThe RNA-Seq analysis provided comprehensive transcriptome information on E.gracilis for the first time, and this information will advance our understanding of this unique organism. The comprehensive analysis indicated that paramylon and wax ester metabolic pathways are regulated at post-transcriptional rather than the transcriptional level in response to anaerobic conditions.

Dataset Information

Comparative Analysis of Strategies for De Novo Transcriptome Assembly in Prokaryotes: Streptomyces clavuligerus as a Case Study.

Publications

Comparative Analysis of Strategies for <i>De Novo</i> Transcriptome Assembly in Prokaryotes: <i>Streptomyces clavuligerus</i> as a Case Study.

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