Difficult. First you should think about what a "change in mRNA concentration" for a particular gene means (should mean). It is relevant if a gene is, say, 2fold induced when allmost other genes are also 2fold induced?

I think it would be wise to use related genes (in terms of the physiological response you are analyzing) and compare the relative expressions of these related genes. This might tell you something about alterations in the regulation of this process.

I agree, it is difficult to find control gene, usually even set of control genes are recommended to normalize real time pcr. Especially it is a problem for complex genomes. Why until now there is not much alternative methods to bypass this problem, or this methods are not widely used? I have attached an interesting publication...

Thanks for the responses and the paper, Angelika. To answer Jochen's question: I'm looking for a housekeeping gene (alpha tubulin, beta actin, EF1alph, etc) that ensures the cDNA levels are the same across samples. It would seem to me, if there is change in mRNA concentration across all genes in the same pattern that might be reflecting different amount of input template or something else biological. I would need a gene that's no affected to know for sure. Alternatively, if the change in mRNA abundance correlated in a dose dependent way that would be suggestive.

To check for your RNA abundance you might think of to make a pool from your each RNA with the same mass (calculated from your conc. values) and divide this pool 1) for the RNA gel 2) for the cDNA Synthesis. I do it so, because in this way I can both check the integrity of my RNA and the correctness of my measurements. By doing so, you can better be sure that the Situation you see on the gel, highly probably resembles what you have with your cDNA. Nevertheless, I use an internal control also.

If you hypothesize that your Treatment would Change global transcriptome, then it is itself contradictory to look for such an internal control also. Although it is under a lot of debate still, you may consider using spike-in technique. Additionally, you may make a thorough search in GEO database of NCBI and look for Treatments, which possibly give you an idea about your Treatment. By analyzing those data (easier to do it with BRB Array Tools or web based microarray Analysis tools) you can see unchanged genes and take them as possible candidates for your internal control.

Good luck!

By the way, 300 ng of total RNA is enough to see the differences on a 1.2% formaldehyde agarose gel.

Looking at related genes, as Jochen suggests, is a good idea. If you think that RNA polymerase II-mediated transcription is affected globally, then you could try to normalise levels to transcripts produced by one of the other polymerases, e.g. U6, see http://www.pnas.org/content/83/22/8575.full.pdf.

Thanks Muammer and Daimark,

That was useful advice you provided. I hadn't considered looking at alternate polymerases or checking cDNA, which I eventually did using a nanodrop.

Normally the transcription factors are chosen for these kind of studies. I recommend you choose 4 from literature basedand check the change under control and your problem condition by qPCR. The DDCT most be almost equal for both conditions if your houskeeping is the correct.