bacterial housekeeping genes

Jim meador at cts.com
Sat Feb 10 12:27:28 EST 1996


Michael,

GAPDH is not suitable for a "housekeeping" gene if your comparing samples
from different cells, whether they are the same cell line but different
samples or are different cell lines/strains. Levels of GAPDH apparently
change with the cell cycle. I don't have references handy, but a search of
Entrez would certainly find some. However, I am basing my statement on my
own observations using RNase protection results. Actin also has its
problems although it is used about as often as GAPDH. 

I, personally, think the bottom line is that it depends on how your
experiment is set up. If you must compare different cell samples, as is
usual, your going to have a hard time. However, you might as well just go
with actin and/or GAPDH, since everyone else does and the reviewers seem
to accept it.  They may can it for a stupid reason, but they probably
won't can it for a good reason like this. Just try to be as careful as you
can, and do at least 2-3 repeats to see what the variability is.

One possible gene to try would be beta-2-macroglobulin, since it seems to
be constitutively expressed and is expressed in most every cell in the
body. Whether it is affected by cell cycle or not, I don't know. Maybe
someone else has looked at this already?

Good luck,
Jim



In article <mcgorry-0802961338000001 at quadra950.pathology.pitt.edu>,
mcgorry at med.pitt.edu (Michael C. Gorry) wrote:

I have been looking for a bacterial housekeeping gene that has
a high level of expression.  Would GADPH be suitable?  We are
looking for any gene that would have a high mRNA level within
a cell.  I haven't been able to find anything published that discusses 
expression rates within cells.  There was, however, a mention
of using GADPH for an RT-PCR quantitative control.  Is anyone
aware of other "controls" that have been used to quantitate PCR
reactions?  I would appreciate any advice.

Thanks

-- 
Michael Gorry
mcgorry at med.pitt.edu



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