Nucleic Acids Research, Vol 27, Issue 19 3821-3835, Copyright © 1999 by Oxford University Press
CS Richmond, JD Glasner, R Mau, H Jin and FR Blattner
We have established high resolution methods for global monitoring of gene
expression in Escherichia coli. Hybridization of radiolabeled cDNA to spot
blots on nylon membranes was compared to hybridization of
fluorescently-labeled cDNA to glass microarrays for efficiency and
reproducibility. A complete set of PCR primers was created for all 4290
annotated open reading frames (ORFs) from the complete genome sequence of
E.coli K-12 (MG1655). Glass- and nylon-based arrays of PCR products were
prepared and used to assess global changes in gene expression. Full-length
coding sequences for array printing were generated by two- step PCR
amplification. In this study we measured changes in RNA levels after
exposure to heat shock and following treatment with isopropyl-
beta-D-thiogalactopyranoside (IPTG). Both radioactive and fluorescence-
based methods showed comparable results. Treatment with IPTG resulted in
high level induction of the lacZYA and melAB operons. Following heat shock
treatment 119 genes were shown to have significantly altered expression
levels, including 35 previously uncharacterized ORFs and most genes of the
heat shock stimulon. Analysis of spot intensities from hybridization to
replicate arrays identified sets of genes with signals consistently above
background suggesting that at least 25% of genes were expressed at
detectable levels during growth in rich media.
ARTICLES
Genome-wide expression profiling in Escherichia coli K-12
Laboratory of Genetics, University of Wisconsin, Madison, WI 53706, USA. craig@genetics.wisc.edu
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