competitive PCR

David R. Johnson david.johnson at yale.edu
Wed Mar 22 17:06:27 EST 2000


Dear Dr. Depoortere,

Do you know why your internal standard RNAs failed?  When you reverse
transcribe, do you prime with random oligos or oligo-dT?  If you used
oligo-dT, are you sure that the internal standard RNAs included a poly-A
sequence?  I was surprised to find that an internal standard RNA with a 15
nucleotide poly A sequence was apparently NOT efficiently primed by oligo-dT,
although I have not yet determined the exact point of failure (hence these
questions to you!).

Regards,
Dave

"Dr. Inge Depoortere" wrote:

> Hi,
> Making internal standard RNAs always failed therefore I have established a
> competitive PCR by adding increasing amounts of a cDNA competitor to  my
> cDNA sample. Although the cDNA competitor has the advantage that it is more
> stable than adding an internal RNA, it has the disadvantage that I can not
> control for the RT. Is there any way I can still control for this step more
> or less.
> Kind regards
> Inge

--
David R. Johnson, Ph.D.
Research Scientist, Department of Pathology
454 BCMM, 295 Congress Avenue
Yale Medical School
New Haven, CT 06510 USA
Tel.: 203/737-2298
Fax: 203/737-2293






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