Help!! transcription of Antisense strand

Peter Ashby p.r.ashby at
Sat Jul 20 09:57:15 EST 2002

In article <20020718040315.25047.qmail at>,
 kunika_jain at wrote:

> Dear All ,
> I have a conceptual problem. I want to produce a PCR  template for 
> transcription of Antisense strand.
> Please verify if my understanding is right, if I say:
> 1. The promoter sequence will be added to the 5' end of the downstream or 
> antisense primer.  If yes then what will be the status of the stop codon that 
> is present as the very last codon of my ORF?? 

it will no longer be the stop codon since you are transcriping the 
antisense, or non coding strand.

> 2. What will be the significance of the added tag that comes after the stop 
> codon.

Since it usually forms only  a very small percentage of the probe, very 
> 2. Is it the right approach?

Yes, though I have found I get better efficiency with some sequence 
5prime of the polymerase promoter.
> 3. In this case do I need to have a start codon in my promoter tag it self.

No. I assume you want to make antisense for a probe. In this case start 
and stop codons are meaningless since you are only making rna not 
translating it into protein.


Peter Ashby
Wellcome Trust Biocentre
University of Dundee, Scotland
Reverse the Spam and remove to email me.

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