hygromycin and arabidopsis

SVemuri12 from gmail.com via methods%40net.bio.net (by SVemuri12 from gmail.com)
Tue Oct 30 17:45:39 EST 2007


I am working with wildtype and transformed lines of Arabidopsis and am
using hygromycin as an antibiotic for selection. On my plates, the
transformed line show good hygromycin resistance, growing into full
green healthy plants with roots. The wildtype line shows good Hyg
sensitivity. So, I knew my antibiotic concentration was working.

Then, I ran the same lines on PCR using Hyg primers. I grew up WT line
separately on soil since they would never survive the Hyg antibiotic.
But I took the transformed lines striaght from Hyg plates and
transferred the ones that survived the antibiotic right to soil. I
prepared genomic DNA after the lines grew old enough.

Now, the PCR shows products in both WT and transformed line. The
product size is the same on both lines, about 500bp. I tested all my
reagants & ran various gels with different annealing temps. But I get
the same products every single time. Does anyone know what this could
mean to see PCR product on the WT line when this line has previously
not shown to be resistant to hygromycin on the Hyg plates? is this
supposed to happen?

Thanks.



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