Biology Reference
In-Depth Information
CHAPTER
analysis
6
Ulrike M ¨ der
*,1
, Pierre Nicolas
{
⁎
Ernst-Moritz-Arndt-University Greifswald, Interfaculty Institute for Genetics
and Functional Genomics, Greifswald, Germany
{
INRA, UR1077 Mathe´matique Informatique et Ge´nome (MIG), Domaine
de Vilvert, Jouy-en-Josas, France
1
Corresponding author. e-mail address: ulrike.maeder@uni-greifswald.de
Array-based approaches
to bacterial transcriptome
1
INTRODUCTION
Systems level approaches to understand bacterial metabolism and its regulation in
response to environmental factors require quantitative, time-resolved data on the
abundance of the cellular components, including mRNAs, proteins, and metabolites.
In bacteria, changes in transcript levels are a key factor in the adaptive response to
changes in environmental conditions. Thus, the analysis of the transcriptome had
become a particularly important part of functional genomics and, in the systems biol-
ogy era, of multi-omics studies aimed at an integrated analysis of the multiple layers
of biological information (
Zhang
et al.
, 2010
). Recent examples include multi-
laboratory efforts to analyse adaptations of metabolism to genetic perturbation in
yeast (
Canelas
et al.
, 2010
) and to environmental changes in the Gram-positive
model bacterium
Bacillus subtilis
(
Buescher
et al.
, 2012
).
Measuring amounts of transcripts at a biologically relevant level of precision
is, at the time of writing, an easier method for acquiring data on cell physiology
than any other systems-level approach. Established techniques for measuring
mRNA abundance such as Northern blotting and real-time (quantitative) PCR
allow for the analysis of limited numbers of genes (
Bustin
et al.
,2005
). However,
with the introduction of methods for genome-wide expression analysis, in partic-
ular, serial analysis of gene expression (
Velculescu
et al.
, 1995
) and DNA micro-
arrays (
Schena
et al.
, 1995
), it became possible to determine the abundance of
thousands of mRNA species simultaneously. In the case of DNA microarrays,
gene-specific probes are attached to a solid surface by employing different array
production technologies. For the analysis, experimental samples are fluorescently
labelled and hybridized to the array. After fluorescence detection by means of a
laser scanning densitometer, the amount of target hybridized to each oligonucle-
otide probe can be quantified from the obtained image. Over the past 15 years,
