rt-pcr problems

Jose M. Bautista jbautist at hgmp.mrc.ac.uk
Tue Jan 2 11:18:22 EST 1996



On 1 Jan 1996, William Sun wrote:

> Hello all,
> 
> I have been trying to amplify messages using RT-PCR, but I don't get any
> bands after the PCR.  I failed to see bands even with primers from 
> Clontech designed to amplify G3PDH.  The protocol that came with the
> primers recommended that I use oligo-dT or random primers during the RT
> step, but I just used the 3' PCR primer.  Has anyone out there have
> success with RT-PCR without using oligo-dT or random priming?  I don't 
> understand why random or oligo-dT priming would give better results.
> 
> -William

How long did you incubate the RT reaction? If less than 1 hour here you
have your trouble: not enough lenght extension of the 1st strand to
amplify by PCR with the second primer.

Hope it help. If not, please, give more details.

Jose




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