mRNA expression in T-DNA insertion mutant - Arabidopsis (Aug/05/2008 )
Hi all,
I have a question about the mRNA in T-DNA insertion mutant. I did real-time PCR recently. I tested the mRNA expression of a gene in its T-DNA insertion mutant. To my surprise, the mRNA level is higher than col-0 wild type. Many paper shows that the mRNA of mutant should be degraded. Does anyone also has this problem?
Can the T-DNA mess up the degradation of inserted mRNA and make inserted mRNA accumulate in Arabidopsis?
Thanks a lot.
-ThomasYang-
QUOTE (ThomasYang @ Aug 5 2008, 09:14 AM)
Hi all,
I have a question about the mRNA in T-DNA insertion mutant. I did real-time PCR recently. I tested the mRNA expression of a gene in its T-DNA insertion mutant. To my surprise, the mRNA level is higher than col-0 wild type. Many paper shows that the mRNA of mutant should be degraded. Does anyone also has this problem?
Can the T-DNA mess up the degradation of inserted mRNA and make inserted mRNA accumulate in Arabidopsis?
Thanks a lot.
I have a question about the mRNA in T-DNA insertion mutant. I did real-time PCR recently. I tested the mRNA expression of a gene in its T-DNA insertion mutant. To my surprise, the mRNA level is higher than col-0 wild type. Many paper shows that the mRNA of mutant should be degraded. Does anyone also has this problem?
Can the T-DNA mess up the degradation of inserted mRNA and make inserted mRNA accumulate in Arabidopsis?
Thanks a lot.
In most cases your mRNA should be degraded, but I suppose there are those rare cases where something funny can happen. It would also depend on the position of your T-DNA insertion. If it's in a promoter or intron, one might imagine a situation where it could have knocked out regulatory sequences. Two things that might help you decide what's going on: 1. confirm that your real time PCR is genuine (make sure that you include a negative control so that you're not amplifying genomic DNA). 2. Perform a northern blot to look at the actual transcripts. Of course if you can do a western blot, that would be even better.
-smu2-