GAL4-AD fusion protein detection/ pGAD-424

Frederik Börnke ricky_boernke at gmx.net
Wed Sep 20 07:02:47 EST 2000


Dear collegues,

my proteins interacts with different 14-3-3 proteins in an 
isoform specific manor in that interaction strength varies
between isoforms. Now I'd like to rule out different
expression levels of my proteins. Western blot is obviously
the method of choice. However, I have my proteins expressed
from pGAD-424 which comprises a truncated ADH promoter 
driving only low expression levels. Mostlikely to low
to detect in a western blot (al least that is what I hear).
So, is there any protocol yielding enough protein to
detect. Or will a commercial lysis solution like Yeast PER
by Pierce help? And what is the best GAL4-AD antibody?
Are there any other methods to ensure comparable expression
levels a referee would accept?

TIA
Ricky


*******************************************************************

Frederik Boernke
Research Group of  Molecular Plant Physiology
Institute for Plant Genetics and Crop Plant Research (IPK)
Corrensstr. 3
06466 Gatersleben
Tel.  039482 -5 321
Fax. 039482 -5 515
http://www.ipk-gatersleben.de






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