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FUNTIONAL GENOMICS: GENOMES AT WORK - Page 2

© Juan C. Mendible

Now in the old times there only two process to look at proteins. One was Polyacrylamide Gel Electrophoresis. The gel is made with wells in which you put the proteins that you extracted from your tissue of interest. You apply an electric current and due to some technological tricks proteins are separated by size. After staining, the result is a ladder of proteins bands, with the smaller ones at the bottom of the gel. Each band representing a number of different proteins of the same size.

The other technique was 2-D (two dimensional) gel electrophoresis. To do this, from the first gel you cut, the ladder that was of interest to you and put it in another gel. You did some new tricks, applied a current a now the proteins of each band were separated by charge. The result now was gel, imagine a square of gelatin, with hundreds of spots. Each one representing one or more proteins that had the same size, we call it molecular weigh, and electrical charge.

Of course, you did not know to which protein corresponded each dot. But by changing the conditions you used to extract the proteins, you could see some of them appearing and/or disappearing, which would tell you how those conditions affected the protein expression pattern of your tissue. You could also compare the expression patterns of different tissues, etc.

With time we learned how to transfer the results of each type of gel to specialized kinds of papers or filters. And we also knew how to purify proteins and make antibodies against them by injecting them into rabbits or other animals. Since antibodies specifically react with their antigens, we reacted those filters with somehow labeled antibodies and so we were able to see if the pattern of expression of our tissue changes according to the conditions we were studying.

But if you stop and think a little, you realize that we are no seeing proteins at work. We are looking at the result of what happened when we changed the conditions of our tissue.

To complicate things a number of the expressed proteins are chemically modified, thus if you are working with a tissue that is capable of expressing 5,000 genes, you can have an unknown number of proteins, say 10,000, doing their jobs at the same time. Doing their jobs means 10,000 molecules colliding between them in an orderly way but that to the untrained eye would look very messy. And believe me, there are at this moment no trained eyes.

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