RT PCR

sbutcher at molbiol.ox.ac.uk sbutcher at molbiol.ox.ac.uk
Tue May 17 12:32:50 EST 1994


MJONES at CRC.AC.UK asked about RT-PCR: 
>1. Do people use a gene specific primer or random hexamers or oligo (dt) or a 
>combination of two or threee as the initial reverse transcription primer.

>2.Does anyone use the thermostable DNA polymerases (eg. Tth) for both RT and 
>PCR amplification.  

Well, I am using the thermostable Perkin Elmer rTth polymerase (together with
their buffers) to amplify short regions from several RNA viruses.
I use a specific reverse primer of 25-27 nucleotides for the reverse
transcription step. So far I have not had any problems with this strategy but 
I have yet to  compare its efficiency directly with reverse transcriptase/TAQ 
strategies. 
Only problem on the horizon is the cost of the enzyme/kit :-(
I have yet to come across anyone else who has used this enzyme so I would be 
very interested in hearing others' experiences with this enzyme.

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Sarah Butcher			SBUTCHER at MOLBIOL.OX.AC.UK
NERC Institute of Virology
Mansfield Road
Oxford UK			usual disclaimer about employer etc.
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