Oligo dT primer - oligo dT primer with non-T priming 3´ or with 5´ (Jul/19/2008 )

Hi there,

I have seen that people use oligo dT with non-T priming 3´ or 5´-V-oligo dT in their RT-PCR reaction. I was wondering how this works and what the beneift is.

Cheers, Siavoush

Hi Siavoush,
I think you mean anchored-oligo(dT) priming for RT-PCR. Using anchored oligo dT leads to binding at the beginning of the poly(A) tail of the mRNA to generate full-length cDNAs and to prevent priming from internal sites of the poly(A) tail. The 5' ends of long mRNAs are often underrepresented; therefore, this priming method is preferred for most applications.
Anchored Oligo(dT) Primers have x thymidine residues and, to ensure binding at the beginning of the mRNA message, one G, C or A residue (the anchor) at the 3' end.
Some companies make the anchor out of 1 G/C/A-residue at the 3' end, others out of 2 (e.g. AA, AT, AC, AG, GA, GC, GT, GG, ...) which is even better.
In the end you have a higher efficiency in your RT-PCR and usually generate longer cDNA fragments.
Hope this answers your question...

Usually it is cheaper to simply order the oligo than to buy it from companies or as part of a kit. Order the oligo
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Thanks for your help.

Siavoush