MadSci Network: Molecular Biology

Re: Why is Gap-dh often used as a housekeeping protein for western blots?

Date: Mon Oct 9 17:48:24 2000
Posted By: Pamela Norton, faculty, Dept. of Medicine, Thomas Jefferson Univ.
Area of science: Molecular Biology
ID: 963245261.Mb

GAPDH (aka glyceraldehyde-3-phosphate dehydrogenase) is one of a number of 
so called housekeeping genes. These genes code for proteins that are 
expressed at a relatively constant rate, both hypothetically (as they are 
thought to be required for cell viability) and experimentally (they tend 
not to vary greatly in amount). 

The value of a protein (or RNA) that is present at a constant level is that 
it provides a reference against which to compare your protein (or RNA) of 
interest. For instance, one dish of cells is treated with a drug, and an 
identical dish is left untreated. Using an antibody to detect protein X by 
Western blot, it appears that drug treatment increases protein X levels 3 
fold. But is this a general increase in protein level? Or is it some sort 
of experimental artifact? The experiment is repeated using an antibody to 
GAPDH; no difference is seen between treated and untreated cells. Taken 
together, the results suggest that there is a specific effect of the drug 
on Protein X levels. In the ideal case, one would always compare test 
protein levels against a control present at a constant amount. In practice, 
nothing turns out to be absolutely constant, and many people normalize to 
loading equal amounts of total protein.

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