On Jun 16, 7:35 am, "Abo-Ogiala, Atef" <Atef.Abo-Ogi... from forst.uni-
> I did the PCR reaction for one of Arabidopsis mutants to verify the T-DNA
> insertion and the results are a bit strange. I mean I should get from the
> first generation of the mutant at least one hetero plant but that is not
> happened. The whole plants were wild type. So it could be there is no T-DNA
> insertion or what? Another question concerning cDNA, I tested the cDNA with
> the specific primers of my gene (Lp+Rp) and also with (Rp+LB) and I got
> results in both reactions, however I got the RNA from wildtype plants. For
> that I have a doubt that cDNA could be contaminated or I have no idea?
>> Best regards
>> Atef Abo-Ogiala
Use the SIGnAL T-DNA Express tool to find the T-DNA orientation for
your mutants. (http://signal.salk.edu/cgi-bin/tdnaexpress). As
recommendation avoid the use of LBb1 primer for Salk mutants, instead
use the LBb1.3 or LBa1 primer. Regarding to your PCR on cDNA just
repeat it but extract RNA from Arabidopsis WT plants grow up by
another person, if no possible grow a new batch of WT plants.
Hope this help!