Short PCR product

aes ali at image.dk
Wed Jan 9 04:12:00 EST 2002


 am not hoping to do absolute real time quantitation, as making a standard
is not possible. However it should be possible to compare to samples if I
can get the PCR working?

> I don't think reliable quantification is possible with degenerate
> primers. The standards will be different from the target and therefore
> amplification efficiency will vary. This doesn't sound like a reliable
>
> method.
>
> With best regards
> Nicolas
>
> -----------------------------------------
> University Hospital Goettingen
> Internal Medicine
> Dept. Klinische Chemie/Clinical Chemistry
> Dr. med. Nicolas von Ahsen
> Robert-Koch-Str. 40
> 37075 Goettingen
> Germany
>
>
> On Tue, 8 Jan 2002 13:48:37 +0000, Duncan Clark <junk@[127.0.0.1]>
> wrote:
>
> >In message <a1egi1$aha$1 at news.net.uni-c.dk>, aes <ali at image.dk> writes
> >on the Tue, 8 Jan 2002
> >>I am going to attempt to do quantitative PCR with degenerate primers, I
can
> >>make the primers so that they should amplify all the genes I want, but
again
> >>the primers end up being 28 bp long with 8 bases (in the middle) having
> >>degenerate composition (R,K and Y). Do you think it is possible to get
these
> >>primers working in a semi quantitative experiment?
> >
> >I would think you are looking for trouble with false priming and/or
> >primer dimers. You would need to verify the product you are seeing is
>





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