Dataset Information

Mapping the RNA-Seq trash bin: unusual transcripts in prokaryotic transcriptome sequencing data.

ABSTRACT: Prokaryotic transcripts constitute almost always uninterrupted intervals when mapped back to the genome. Split reads, i.e., RNA-seq reads consisting of parts that only map to discontiguous loci, are thus disregarded in most analysis pipelines. There are, however, some well-known exceptions, in particular, tRNA splicing and circularized small RNAs in Archaea as well as self-splicing introns. Here, we reanalyze a series of published RNA-seq data sets, screening them specifically for non-contiguously mapping reads. We recover most of the known cases together with several novel archaeal ncRNAs associated with circularized products. In Eubacteria, only a handful of interesting candidates were obtained beyond a few previously described group I and group II introns. Most of the atypically mapping reads do not appear to correspond to well-defined, specifically processed products. Whether this diffuse background is, at least in part, an incidental by-product of prokaryotic RNA processing or whether it consists entirely of technical artifacts of reverse transcription or amplification remains unknown.

SUBMITTER: Doose G

PROVIDER: S-EPMC3849169 | biostudies-literature | 2013 Jul

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Mapping the RNA-Seq trash bin: unusual transcripts in prokaryotic transcriptome sequencing data.

Doose Gero G Alexis Maria M Kirsch Rebecca R Findeiß Sven S Langenberger David D Machné Rainer R Mörl Mario M Hoffmann Steve S Stadler Peter F PF

RNA biology 20130513 7

Prokaryotic transcripts constitute almost always uninterrupted intervals when mapped back to the genome. Split reads, i.e., RNA-seq reads consisting of parts that only map to discontiguous loci, are thus disregarded in most analysis pipelines. There are, however, some well-known exceptions, in particular, tRNA splicing and circularized small RNAs in Archaea as well as self-splicing introns. Here, we reanalyze a series of published RNA-seq data sets, screening them specifically for non-contiguous ...[more]

PMID: 23702463

Similar Datasets

Project description:Background Nowadays, both customers and producers prefer thin-tailed fat sheep. To effectively breed for this phenotype, it is important to identify candidate genes and uncover the genetic mechanism related to tail fat deposition in sheep. Accumulating evidence suggesting that post-transcriptional modification events of precursor-messenger RNA (pre-mRNA), including alternative splicing (AS) and alternative polyadenylation (APA), may regulate tail fat deposition in sheep. Differentially expressed transcripts (DETs) analysis is a way to identify candidate genes related to tail fat deposition. However, due to the technological limitation, post-transcriptional modification events in the tail fat of sheep and DETs between thin-tailed and fat-tailed sheep remains unclear. Methods In the present study, we applied pooled PacBio isoform sequencing (Iso-Seq) to generate transcriptomic data of tail fat tissue from six sheep (three thin-tailed sheep and three fat-tailed sheep). By comparing with reference genome, potential gene loci and novel transcripts were identified. Post-transcriptional modification events, including AS and APA, and lncRNA in sheep tail fat were uncovered using pooled Iso-Seq data. Combining Iso-Seq data with six RNA-sequencing (RNA-Seq) data, DETs between thin- and fat-tailed sheep were identified. Protein protein interaction (PPI) network, Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses were implemented to investigate the potential functions of DETs. Results In the present study, we revealed the transcriptomic complexity of the tail fat of sheep, result in 9,001 potential novel gene loci, 17,834 AS events, 5,791 APA events, and 3,764 lncRNAs. Combining Iso-Seq data with RNA-Seq data, we identified hundreds of DETs between thin- and fat-tailed sheep. Among them, 21 differentially expressed lncRNAs, such as ENSOART00020036299, ENSOART00020033641, ENSOART00020024562, ENSOART00020003848 and 9.53.1 may regulate tail fat deposition. Many novel transcripts were identified as DETs, including 15.527.13 (DGAT2), 13.624.23 (ACSS2), 11.689.28 (ACLY), 11.689.18 (ACLY), 11.689.14 (ACLY), 11.660.12 (ACLY), 22.289.6 (SCD), 22.289.3 (SCD) and 22.289.14 (SCD). Most of the identified DETs have been enriched in GO and KEGG pathways related to extracellular matrix (ECM). Our result revealed the transcriptome complexity and identified many candidate transcripts in tail fat, which could enhance the understanding of molecular mechanisms behind tail fat deposition.

Project description:Anopheles funestus is one of the primary vectors of human malaria, which causes a million deaths each year in sub-Saharan Africa. Few scientific resources are available to facilitate studies of this mosquito species and relatively little is known about its basic biology and evolution, making development and implementation of novel disease control efforts more difficult. The An. funestus genome has not been sequenced, so in order to facilitate genome-scale experimental biology, we have sequenced the adult female transcriptome of An. funestus from a newly founded colony in Burkina Faso, West Africa, using the Illumina GAIIx next generation sequencing platform.We assembled short Illumina reads de novo using a novel approach involving iterative de novo assemblies and "target-based" contig clustering. We then selected a conservative set of 15,527 contigs through comparisons to four Dipteran transcriptomes as well as multiple functional and conserved protein domain databases. Comparison to the Anopheles gambiae immune system identified 339 contigs as putative immune genes, thus identifying a large portion of the immune system that can form the basis for subsequent studies of this important malaria vector. We identified 5,434 1:1 orthologues between An. funestus and An. gambiae and found that among these 1:1 orthologues, the protein sequence of those with putative immune function were significantly more diverged than the transcriptome as a whole. Short read alignments to the contig set revealed almost 367,000 genetic polymorphisms segregating in the An. funestus colony and demonstrated the utility of the assembled transcriptome for use in RNA-seq based measurements of gene expression.We developed a pipeline that makes de novo transcriptome sequencing possible in virtually any organism at a very reasonable cost ($6,300 in sequencing costs in our case). We anticipate that our approach could be used to develop genomic resources in a diversity of systems for which full genome sequence is currently unavailable. Our An. funestus contig set and analytical results provide a valuable resource for future studies in this non-model, but epidemiologically critical, vector insect.

Project description:The advent of next generation sequencing technologies (NGS) has expanded the area of genomic research, offering high coverage and increased sensitivity over older microarray platforms. Although the current cost of next generation sequencing is still exceeding that of microarray approaches, the rapid advances in NGS will likely make it the platform of choice for future research in differential gene expression. Connectivity mapping is a procedure for examining the connections among diseases, genes and drugs by differential gene expression initially based on microarray technology, with which a large collection of compound-induced reference gene expression profiles have been accumulated. In this work, we aim to test the feasibility of incorporating NGS RNA-Seq data into the current connectivity mapping framework by utilizing the microarray based reference profiles and the construction of a differentially expressed gene signature from a NGS dataset. This would allow for the establishment of connections between the NGS gene signature and those microarray reference profiles, alleviating the associated incurring cost of re-creating drug profiles with NGS technology. We examined the connectivity mapping approach on a publicly available NGS dataset with androgen stimulation of LNCaP cells in order to extract candidate compounds that could inhibit the proliferative phenotype of LNCaP cells and to elucidate their potential in a laboratory setting. In addition, we also analyzed an independent microarray dataset of similar experimental settings. We found a high level of concordance between the top compounds identified using the gene signatures from the two datasets. The nicotine derivative cotinine was returned as the top candidate among the overlapping compounds with potential to suppress this proliferative phenotype. Subsequent lab experiments validated this connectivity mapping hit, showing that cotinine inhibits cell proliferation in an androgen dependent manner. Thus the results in this study suggest a promising prospect of integrating NGS data with connectivity mapping.

Dataset Information

Mapping the RNA-Seq trash bin: unusual transcripts in prokaryotic transcriptome sequencing data.

Publications

Mapping the RNA-Seq trash bin: unusual transcripts in prokaryotic transcriptome sequencing data.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets