housekeeping genes for brain tissue - (Sep/29/2008 )
Hi,
do you know any houeskeeping genes specially suited for doing RT-PCR out of brain tissue (we use hippocampus region from cats) apart from beta-actin or GAPDH?
Furthermore I have real basic questions concerningt the definition of the term "housekepping gene" - 1) are the quantities of housekeeping gene-expression different in different cell types and 2) is the quantity of hoousekeeping gene-expression different between cells of the same cell ype dependant on the localisation (e.g. is housekeeping gene-expression in neurons of the hippocampus different from that of neurons in other brain areas?
Many thanks,
Barbara
i would say state-of-the-art procedure is to validate the most suitable reference genes and normalize to the mean expression of them.
this has been already discussed in this forum. just use the search option.
do you know any houeskeeping genes specially suited for doing RT-PCR out of brain tissue (we use hippocampus region from cats) apart from beta-actin or GAPDH?
Furthermore I have real basic questions concerningt the definition of the term "housekepping gene" - 1) are the quantities of housekeeping gene-expression different in different cell types and 2) is the quantity of hoousekeeping gene-expression different between cells of the same cell ype dependant on the localisation (e.g. is housekeeping gene-expression in neurons of the hippocampus different from that of neurons in other brain areas?
Many thanks,
Barbara
There was an article in Biotechniques last year IIRC which mentioned a few. I use 18s and PPIA for retina and superior colliculus. You're going to have to validate them all to your particular assay though.
Hi Barbara,
You may want to try Genevestigator and RefGenes. Genevestigator allows you to enter your gene of interest and look at how this gene is expressed in different tissues, different stages of development, or how it responds to all kinds of conditions. In this way, you can kind of validate existing candidates.
On the other hand, you can also search for new reference genes, which is the better solution in your case. RefGenes is a tool from Genevestigator that allows you to find new reference genes based on a selected set of conditions, such as tissues. Our experience with the proposed genes is very positive and they were always performing better than classical ones such as ACTB and 28S. Here's where you can find these tools:
http://www.genevestigator.com
https://www.genevestigator.ethz.ch/gv/doc/refgenes.jsp
Philip
For my work, I use HPRT and 36B4.
Since there is NO SUCH GENE consistently express among tissues/cell types and among various conditions (I will consider same cell type at different localization as different conditions), I always take time do internal control selection. Thus I can have better idea what gene(s) will be the best for normalization.