Dataset Information

SMRT-Cappable-seq reveals complex operon variants in bacteria.

ABSTRACT: Current methods for genome-wide analysis of gene expression require fragmentation of original transcripts into small fragments for short-read sequencing. In bacteria, the resulting fragmented information hides operon complexity. Additionally, in vivo processing of transcripts confounds the accurate identification of the 5' and 3' ends of operons. Here we develop a methodology called SMRT-Cappable-seq that combines the isolation of un-fragmented primary transcripts with single-molecule long read sequencing. Applied to E. coli, this technology results in an accurate definition of the transcriptome with 34% of known operons from RegulonDB being extended by at least one gene. Furthermore, 40% of transcription termination sites have read-through that alters the gene content of the operons. As a result, most of the bacterial genes are present in multiple operon variants reminiscent of eukaryotic splicing. By providing such granularity in the operon structure, this study represents an important resource for the study of prokaryotic gene network and regulation.

SUBMITTER: Yan B

PROVIDER: S-EPMC6131387 | biostudies-literature | 2018 Sep

REPOSITORIES: biostudies-literature

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SMRT-Cappable-seq reveals complex operon variants in bacteria.

Yan Bo B Boitano Matthew M Clark Tyson A TA Ettwiller Laurence L

Nature communications 20180910 1

Current methods for genome-wide analysis of gene expression require fragmentation of original transcripts into small fragments for short-read sequencing. In bacteria, the resulting fragmented information hides operon complexity. Additionally, in vivo processing of transcripts confounds the accurate identification of the 5' and 3' ends of operons. Here we develop a methodology called SMRT-Cappable-seq that combines the isolation of un-fragmented primary transcripts with single-molecule long read ...[more]

PMID: 30201986

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Project description:BackgroundCardamine violifolia, native to China, is one of the selenium (Se) hyperaccumulators. The mechanism of Se metabolism and tolerance remains unclear, and only limited genetic information is currently available. Therefore, we combined a PacBio single-molecule real-time (SMRT) transcriptome library and the Illumina RNA-seq data of sodium selenate (Na2SeO4)-treated C. violifolia to further reveal the molecular mechanism of Se metabolism.ResultsThe concentrations of the total, inorganic, and organic Se in C. violifolia seedlings significantly increased as the Na2SeO4 treatment concentration increased. From SMRT full-length transcriptome of C. violifolia, we obtained 26,745 annotated nonredundant transcripts, 14,269 simple sequence repeats, 283 alternative splices, and 3407 transcription factors. Fifty-one genes from 134 transcripts were identified to be involved in Se metabolism, including transporter, assimilatory enzyme, and several specific genes. Analysis of Illumina RNA-Seq data showed that a total of 948 differentially expressed genes (DEGs) were filtered from the four groups with Na2SeO4 treatment, among which 11 DEGs were related to Se metabolism. The enrichment analysis of KEGG pathways of all the DEGs showed that they were significantly enriched in five pathways, such as hormone signal transduction and plant-pathogen interaction pathways. Four genes related to Se metabolism, adenosine triphosphate sulfurase 1, adenosine 5'-phosphosulfate reductase 3, cysteine (Cys) desulfurase 1, and serine acetyltransferase 2, were regulated by lncRNAs. Twenty potential hub genes (e.g., sulfate transporter 1;1, Cys synthase, methionine gamma-lyase, and Se-binding protein 1) were screened and identified to play important roles in Se accumulation and tolerance in C. violifolia as concluded by weighted gene correlation network analysis. Based on combinative analysis of expression profiling and annotation of genes as well as Se speciation and concentration in C. violifolia under the treatments with different Na2SeO4 concentrations, a putative Se metabolism and assimilation pathway in C. violifolia was proposed.ConclusionOur data provide abundant information on putative gene transcriptions and pathway involved in Se metabolism of C. violifolia. The findings present a genetic resource and provide novel insights into the mechanism of Se hyperaccumulation in C. violifolia.

Dataset Information

SMRT-Cappable-seq reveals complex operon variants in bacteria.

Publications

SMRT-Cappable-seq reveals complex operon variants in bacteria.

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