Degenerate primers for direct fragment sequencing

Mark nospam at nspam.com
Wed Jun 1 17:24:04 EST 2005


amerotto wrote:
> I am sequencing fragments from Cyperus difformis (Plant, cyperaceae). I
> am using
> degenarate primers in order to isolate the fragments that I want. For
> some
> primers/template combination I had good results, but for another the
> sequence just not
> appear or appear with double peaks. I am using the Qiagen gel cleaning
> kit and the
> Sigma spin column for the cleaning procedures.
> 
> Question: Are degenerate primers a problem for sequencing? If yes, how
> can I compensate
> for that?
> 
> Thanks
> 
> Aldo
> 
> 
> Aldo Merotto
> 
you could clone your PCR products in a plasmid that has sequences 
for (for example) T7 and T3 universal primers, and then just use 
those.



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