PCR'd DNA vs RNA probes for northerns

Vivian Miao vmiao at oregon.uoregon.edu
Sat Oct 30 14:26:38 EST 1993

In article <2au7g6$ojo at usenet.INS.CWRU.Edu>, Harry Menegay wrote:

> 	We are currently making RNA probes to probe northern blots, however we are
> considering going to DNA probes made from single pimer PCR.  Has anyone tried one
> versus the other? Is background better or worse? Is the signal decreased?

Hi Harry,  I don't know if this addresses your question about background,
but my "normal" probes for Northerns are made by the random hex method off
a gel isolated fragment or off a PCR product, so presumably both strands
are there.  I know half the probe is going to waste, but the signal is
fine, and with respect to your concern - there isn't any background
problem.  I work with Neurospora crassa, so maybe the small genome (43
megabases) has something to do with why I can get away with this.  

I also have a protocol for making DNA probes by "single primer PCR" (linear
amplification) which I'll be glad to post or email if it is of interest.

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