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J Bacteriol. 2015 Jan 1;197(1):18-28. doi: 10.1128/JB.02096-14. Epub 2014 Sep 29.

Global transcriptional start site mapping using differential RNA sequencing reveals novel antisense RNAs in Escherichia coli.

Author information

  • 1Cell Biology and Metabolism Program, Eunice Kennedy Shriver National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, Maryland, USA.
  • 2Research Center for Infectious Diseases (ZINF), University of Würzburg, Würzburg, Germany.
  • 3Cell Biology and Metabolism Program, Eunice Kennedy Shriver National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, Maryland, USA Research Center for Infectious Diseases (ZINF), University of Würzburg, Würzburg, Germany.
  • 4Center for Bioinformatics Tübingen (ZBIT), University of Tübingen, Tübingen, Germany.
  • 5Cell Biology and Metabolism Program, Eunice Kennedy Shriver National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, Maryland, USA Research Center for Infectious Diseases (ZINF), University of Würzburg, Würzburg, Germany cynthia.sharma@uni-wuerzburg.de storzg@mail.nih.gov.
  • 6Cell Biology and Metabolism Program, Eunice Kennedy Shriver National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, Maryland, USA cynthia.sharma@uni-wuerzburg.de storzg@mail.nih.gov.

Abstract

While the model organism Escherichia coli has been the subject of intense study for decades, the full complement of its RNAs is only now being examined. Here we describe a survey of the E. coli transcriptome carried out using a differential RNA sequencing (dRNA-seq) approach, which can distinguish between primary and processed transcripts, and an automated prediction algorithm for transcriptional start sites (TSS). With the criterion of expression under at least one of three growth conditions examined, we predicted 14,868 TSS candidates, including 5,574 internal to annotated genes (iTSS) and 5,495 TSS corresponding to potential antisense RNAs (asRNAs). We examined expression of 14 candidate asRNAs by Northern analysis using RNA from wild-type E. coli and from strains defective for RNases III and E, two RNases reported to be involved in asRNA processing. Interestingly, nine asRNAs detected as distinct bands by Northern analysis were differentially affected by the rnc and rne mutations. We also compared our asRNA candidates with previously published asRNA annotations from RNA-seq data and discuss the challenges associated with these cross-comparisons. Our global transcriptional start site map represents a valuable resource for identification of transcription start sites, promoters, and novel transcripts in E. coli and is easily accessible, together with the cDNA coverage plots, in an online genome browser.

PMID:
25266388
PMCID:
PMC4288677
DOI:
10.1128/JB.02096-14
[PubMed - indexed for MEDLINE]
Free PMC Article
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