attaching t7 promoter sequence to template primer - (Jan/10/2007 )
I wanted to design primers with the T7 promoter sequence so that i can transcribe and produce RNA for a riboprobe.
if anyone can help me on which primer do I add the T7 promoter sequence? i think it's the forward.
If you're interested to look at mRNA expression in in situ hybridization, you'll have to have T7 in the same strand as the anti-sense. I.e. you have to transcribe anti-sense RNA (labelled) that'll hybridize to mRNA of interest.
If you're interested to look at the dsDNA (either gDNA, vector etc), both strands can be use and the T7 location does not matter.
NOTE: Some vectors have T7 and T3 around the MCS. If you've cloned your gene in there, you could opt for T7 transcription or T3 transcription to get the RNA you're interested.
Hope this helped.