distinguish protein expression from high homology - (Jun/16/2009 )
Hello,
I'm working on transgenic mice. Now i have trouble in detecting overexpression of Tg animal. Since the endogenous and overexpressed cDNA are 98% homology in protein sequence, there is no anitibody available to separate them. I've been trying fluorescent western blot which might be able to distinguish 30% difference, but still not that accurate.
Does anyone know other methods to separate them? Or any modification in transgene which could be distinguished from endogenous one?
Many thanks!
YC
-harmodrie-
harmodrie on Jun 16 2009, 02:31 AM said:
Hello,
I'm working on transgenic mice. Now i have trouble in detecting overexpression of Tg animal. Since the endogenous and overexpressed cDNA are 98% homology in protein sequence, there is no anitibody available to separate them. I've been trying fluorescent western blot which might be able to distinguish 30% difference, but still not that accurate.
Does anyone know other methods to separate them? Or any modification in transgene which could be distinguished from endogenous one?
Many thanks!
YC
I'm working on transgenic mice. Now i have trouble in detecting overexpression of Tg animal. Since the endogenous and overexpressed cDNA are 98% homology in protein sequence, there is no anitibody available to separate them. I've been trying fluorescent western blot which might be able to distinguish 30% difference, but still not that accurate.
Does anyone know other methods to separate them? Or any modification in transgene which could be distinguished from endogenous one?
Many thanks!
YC
Hi YC,
just a thought. why don't you add some tag, myc, GST or GFP et al, to your cDNA so that you can tell from the endogenous protein with the antibody or fluorescence to the tag region.
D
-ixsix-