You are interested in understanding how the brain works, and are using the fruit fly Drosophila as a model system to study brain development. You perform a microarray analysis to try to determine genes expressed in the fly brain
For your microarray experiment, you first prepare cDNA from fly brains and label it with a red fluorochrome. Then you isolate cDNA from whole flies and label it with a green fluorochrome. Next you hybridize these cDNA populations to a microarray containing the Drosophila genes. From this you obtain a list of genes that are specifically enriched in the brain (those that show up as a red spot on the microarray).
You are disappointed because your favorite fly gene, tubby, does not appear on this list, even though you have repeated the microarray experiment 10 times and did not encounter any technical difficulties. The reason you thought tubby would appear on this list is that you believe tubby is important for brain development, because flies mutant in tubby have no brains. Not to be discouraged, you perform in situ analysis using the tubby DNA as a probe, and see that it is expressed at high levels in the fly brain of normal flies but not expressed in animals lacking the tubby gene.
Why do you think tubby did not show up as a gene specifically enriched in the brain in your microarray experiment?
The tubby gene is expressed in all tissues, including the brain. A red spot on a microarray is indicative of a difference in gene expression between the two RNA samples being compared. You may expect in this experiment that the tubby gene would be a yellow spot (a gene expressed at equal levels in both samples). An in situ experiment looks at the RNA level directly in the tissue of interest, which is why in this case you see ample levels of tubby RNA.
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