endogenous GUS activity

Daniel PERAZZA perazza at bio.grenet.fr
Wed Oct 2 12:37:07 EST 1996


Dear arabinetters,

I am analysing GUS expression driven by a weak promoter in transgenic
plants using fluorimetric assay, and I am facing problems. I can only
extract small amounts of total protein (20-40 micrograms) from young
developing leaves (tissue where the promoter is active in transgenic plants
as shown by histochemical X-gluc staining). When I compare enzymatic
activities from transgenic vs wt control plants, I cannot see differences.
Apparently, in my experimental conditions (pH 7.0, 37=B0C) wt protein
extracts are able to hydrolyse small amounts of MUG (substrate) so that I
am unable to distinguish transgenic from wt (wt GUS-like specific activity
0.6 pmol/h/microgram of protein).
Does anybody have a method to decrease endogenous level of GUS-like
activity that I observe at pH 7.
Any comments or ideas will be welcome. Thank you in advance for your respons=
es.
Daniel




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        PERAZZA Daniel

address:Laboratoire de Genetique Moleculaire des Plantes - UMR/CNRS 5575
        Universite Joseph Fourier
        BP 53
        F38041 GRENOBLE cedex 09
Tel:    (33) 76-51-48-92
=46ax:    (33) 76-51-43-36
Email:  perazza at bio.grenet.fr
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