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[Arabidopsis] Protoplast transformation and HA antibody.

Hicham Zegzouti hichamz at mcdb.ucla.edu
Tue Jan 31 13:58:53 EST 2006


Dear Arabidopsis community,

I am trying to express an HA-tagged protein (50KD) in protoplast 
using PEG transformation. When I run a western after 18-24 hours 
incubation, I don't see any differentially present band corresponding 
to 50 KD present in my transformed protoplast lane. All the bands 
that were present on the gel are detected non specifically by the HA 
antibody I am using (Covance). When I use GFP as a control for 
transformation, the GFP antibody recognizes the GFP band showing that 
the transformation protocol I am using works at least for a 30KD 
protein. I am thinking that the HA antibody (Covance) I am using may 
not be good.

I would be very greatful if you could tell me about any experience 
you have with HA tagged proteins and what antibody you have used 
successfully for westerns and immunoprecipitations from plant, plant 
cells or protoplasts.

Thank you from your help

Hicham Zegzouti, PhD
UCLA

-- 
**************************************************************************
Hicham ZEGZOUTI, Ph.D
UCLA
Department of Molecular, Cell and Developmental Biology
Christensen lab
621 Charles E. Young Drive, South
P.O. Box 951606
Los Angeles, CA 90095-1606
Phone/Fax: 1 310 794 7096
email: hichamz at mcdb.ucla.edu
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