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Today I looked at protein expression of a flowering time experiment. I evaluated protein expression by western blotting. This is a fairly straight-forward concept that allows you to see relative amounts of protein. I began by extracting total protein from plants that I was interested in evaluating protein expression/accumulation. This process starts by clipping a single leaf from a plant and grinding the leaf in a tube with an appropriate amount of extraction buffer. The extraction buffer allows the protein to go into solution with a cold centrifugation step. After centrifugation, everything except total protein is pelleted to the bottom of the tube. I then remove this top portion, called the supernatant. This protein rich extract is transferred to a tube containing loading dye, and heated at 95 degrees for 5 minutes. This heating step denatures the protein, or linearizes it, in other words. We are now ready to load our proteins into a poly-acrylamide gel that utilizes charge to push the proteins through the gel. The different proteins are separated by size, larger proteins remain higher up on the gel, smaller ones drift further down.