NDLI: Immobilization of Escherichia coli RNA Polymerase and Location of Binding Sites by Use of Chromatin Immunoprecipitation and Microarrays†

Content Provider	PubMed Central
Author	Herring, Christopher D. Raffaelle, Marni Allen, Timothy E. Kanin, Elenita I. Landick, Robert Ansari, Aseem Z. Palsson, Bernhard Ø.
Copyright Year	2005
Abstract	The genome-wide location of RNA polymerase binding sites was determined in Escherichia coli using chromatin immunoprecipitation and microarrays (chIP-chip). Cross-linked chromatin was isolated in triplicate from rifampin-treated cells, and DNA bound to RNA polymerase was precipitated with an antibody specific for the β′ subunit. The DNA was amplified and hybridized to “tiled” oligonucleotide microarrays representing the whole genome at 25-bp resolution. A total of 1,139 binding sites were detected and evaluated by comparison to gene expression data from identical conditions and to 961 promoters previously identified by established methods. Of the detected binding sites, 418 were located within 1,000 bp of a known promoter, leaving 721 previously unknown RNA polymerase binding sites. Within 200 bp, we were able to detect 51% (189/368) of the known σ70-specific promoters occurring upstream of an expressed open reading frame and 74% (273/368) within 1,000 bp. Conversely, many known promoters were not detected by chIP-chip, leading to an estimated 26% negative-detection rate. Most of the detected binding sites could be associated with expressed transcription units, but 299 binding sites occurred near inactive transcription units. This map of RNA polymerase binding sites represents a foundation for studies of transcription factors in E. coli and an important evaluation of the chIP-chip technique.
Related Links	http://dx.doi.org/10.1128/jb.187.17.6166-6174.2005
Ending Page	6174
Page Count	9
Starting Page	6166
File Format	PDF
ISSN	00219193
e-ISSN	10985530
Journal	Journal of Bacteriology
Issue Number	17
Volume Number	187
Language	English
Publisher	American Society for Microbiology
Publisher Date	2005-09-01
Access Restriction	Open
Rights Holder	American Society for Microbiology
Subject Keyword	Molecular Biology Microbiology Research in Higher Education
Content Type	Text
Resource Type	Article
Subject	Molecular Biology Microbiology

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