Competing plasmids in transfections, more details

John Ladasky ladasky at my-deja.com
Thu Mar 21 10:51:10 EST 2002


peter.cherepanov at uz.kuleuven.ac.be ("Peter Cherepanov") wrote in message news:<000901c1d0c0$355efb80$9293a8c0 at uz.kuleuven.ac.be>...
> I just hope that it is not just YFP or CFP folding that is disturbed when
> the complex is formed...

Let me remind you that I have tried two entirely different CFP fusion
protein constructs.  I have protein X fused to YFP.  I have tried
protein Y fused to CFP, and I have also tried protein X fused to CFP. 
In both cases I see the negative correlation between CFP and YFP.  So
for my findings to be explained by  disruption of folding when CFP and
YFP are present, this disruption of folding would have to occur under
two very different conditions.

> I never worked with other FP then GFP, but for some variants of GFP they
> recommened to low down the temperature (30oC), so that it is folding better.
> May be YFP or CFP are less structurally stable then eGFP?

If I express either a CFP protein or the YFP protein individually at
37C, I have no problem.  I only have trouble when I try to express
both of them together.  Still, I will look in to the thermal
stabilities of CFP and YFP as you suggest.  Maybe I can improve my
odds a bit further by reducing the incubator temperature.

Thanks again!

--
John J. Ladasky Jr., Ph.D.
Department of Biology
Johns Hopkins University
Baltimore, MD 21218
USA
Earth




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