what's the best probe to quantitate RNA in Northern blot????

Helge Weissig helgew at ljcrf.edu
Thu Jan 6 14:32:44 EST 1994


>I am studying expression of a mammalian gene at transcription level, under
>different nutritional condition.  I used actin as a probe to quantitate RNA,
>to ensure equal loading in my Northern.  But I didn't have good luck with
>actin
>probe.  So I switched to GAPDH probe.  My bands looked nicer, but since GAPDH
>is also under hormonal control, I can't use it as a quanitative marker either.
>Does anyone have experience with ribosomal RNA (15S?) probe in this regard?
>I will appreciate if you can share your experience here.
>
>Why not just use 10 or 20 ug of total RNA in each lane?  Maybe you ask.
>Yaaaaaa.  I like to hear your logic too.
>
>michael
>mman at molbio.cbs.umn.edu

I had the same experience. When comparing actin and GAPDH it got even more
confusing.... right now I am using L32-4A as a standard. The message for
this ribosomal protein seems to be fairly unregulated (at least in the
cells and tissues I am using). You might want to look into Dudov and Perry,
Cell 37: 457 (1984)

as for the rRNA probe, i have also seen people using these, but that seems
to me as good as an EtBr stained gel... :)

my OD260 = 0.02 worth

good luck,
helge







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