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Concentration of RNA in the cell.

Colin Rasmussen colin at pombe.usask.ca
Fri Jan 21 14:04:13 EST 2000



Alex Harvey wrote:

> Does anyone know the molar concentration of some specific RNAs in the cell,
> just as examples? It could be any cell or any RNA. Also if one knew the
> volume and copy number, how would one make the calculation? For instance, if
> there are 1000 copies of the p53 mRNA in a HeLa cell, what is the
> concentration in nano or pico (or atto?) moles? One could assume (perhaps
> incorrectly) that the mRNA is evenly distributed throughout the cell.

You can do copy number estimates by hybridization with a known sample as your
reference.  One way I've seen this done is to put a known amount of cDNA
containing your sequence on the blot, along with a serial dilution series of
RNA.  Hybridize as usual then compare signals.  This will give you the amount
of your specific mRNA per ug of RNA, or if you load based on cell number per
"x" cells.  If you want concentration per se, you'll then need to take the
amount per cell and divide by the estimated volume.  Easiest way to do this is
to trypsinize cells so they round up, then measure the average diameter with a
micrometer.  Also, some Coulter counters will give you volume information as
well, but you'll need to compare to a known particle size to do that.

Colin

>
>
> Thanks, Alex





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