Protoplast transformation and HA antibody.

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

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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: [Only registered users see links. ]
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