Help!! transcription of Antisense strand
p.r.ashby at MAPS.dundee.ac.uk
Sat Jul 20 09:57:15 EST 2002
In article <20020718040315.25047.qmail at ww02.hostica.com>,
kunika_jain at rediffmail.com 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
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.
Wellcome Trust Biocentre
University of Dundee, Scotland
Reverse the Spam and remove to email me.
More information about the Methods