trouble with RT-PCR

Dag Rune Gjellesvik dagrg_nospam at online.no
Mon Mar 27 05:57:32 EST 2006


Hi,
Maybe your "specific" band comes from a PCR product carry-over
contamination?

Dag Rune Gjellesvik


"Sanni" <Leporis at web.de> skrev i melding
news:1143138712.361791.112850 at v46g2000cwv.googlegroups.com...
> Hi!
> I do RT-PCR with poly dT primers and subsequent PCR from cDNA with
> specific primer pairs. I usually try to analyze the sequence and use
> intron spanning primers. So normally I should be able to distinguish
> between the specific band generated from the cDNA and the larger band
> from contaminating genomic DNA! But if I perform a control RT reaction
> without reverse transcriptase I also see the two bands in the PCR
> reaction of this control ! So how could this "specific" band appear
> without producing cDNA??? I also checked the sequence for retroposons,
> but could not find any! Is it somehow  possible that the specific PCR
> primers could bind to anbundantly expressed mRNAs???
> Any hints or tips would be appreciated ;-)
>
> Best wishes, Sanni
>




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