Functional
Genomics in C. elegans 1:
Microarray Analysis
of Gene Expression
(1: Microarray analysis 2: Expression analysis using reporters 3: RNAi)
With the sequenced genome available, and the extensive EST dataset
from the Kohara laboratory it is now possible to contemplate research
approaches that address the whole genome. One such approach is the
use of arrays of expressed DNA segments as hybridisation targets to
examine gene expression levels. There is an excellent description of
the technology behind the experiments and analysis of microarray experiments at http://www.cs.washington.edu/homes/jbuhler/research/array/experiment.html
This collection of DNA fragments can be printed in dense arrays on
glass slides, and the slide used for hybridisation assays.
Next, two (or more) probes are generated from mRNA from the two
strains being compared. [Its also possible to compare stages, or
differently treated nematodes (drug + vs drug-, for example)]. The
probes are labeled using fluorescently tagged nucleotides. These are
excited by a laser, and their fluorescence read by a tuned
spectrophotometer (much as an autoomated sequencing gel is read).
For example, here is a set of data from an experiment examining
differences between two different nematode strains. One was N2 wild
type, the other was mutant in the let-60 gene. The
let-60 gene encodes a Ras homologue involved in many cell fate
determination decisions in the embryo, and also in processes in later
development, such as vulva formation. The image is from the Kim lab
web site.
