RT-PCR with cultured neuronal cells

Patrick Wefstaedt zeraldas_riese at freenet.de
Fri Jun 11 17:17:21 EST 2004


Hi,
I´m trying to investigate the effects of  Neurotrophic Factors on
neuronal cells from the inner ear (SGC). The cells are kept in culture
for 72h and were harvested afterwards.  In order to determin the
expression levels of various receptors RT-PCR is carried out. As
qualitative proof everything is working fine. My question to you is
wether there is a possibility to try out semiquantitative experiments,
too. The problem is that the RNA from the neuronal cells is decreasing
(because of decreasing number of cells) while the total RNA amount is
increasing as a reason of fibroblast growth. This means that
normalization against a “normal” housekeeping gene e.g.
GAPDH is not possible. Does anybody know about a gene which could be
taken as a “neuronal” housekeeping gene and which is not
regulated after exposition to growth factrors or serum? Do you have
other ideas?

Thanks a lot!

Patrick
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