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Protocol to detect cmyc tagged proteins in plants

daemon at net.bio.net daemon at net.bio.net
Thu Feb 13 11:56:10 EST 1997


Dear Netters,

Does anyone have a working protein extraction/Western blot protocol to
detect cmyc-tagged proteins in plants?

We have tagged two distinct proteins with the 13-amino acid cmyc epitope.
We have then transformed tobacco plants with expression constructs (35S
promoter) carrying these two clones and are now trying to detect the tagged
proteins in whole protein extracts from leaves by Western blots using the
9E-10 cmyc monoclonal antibody which is commercially available in Germany
from Boehringer Mannheim.

Our results are: we cannot detect ANY cross hybridizing bands at all.  Not
even the famous background bands many times mentioned in the literature.

It could well be that the proteins are not being made...  Does any one have
a positive control, a good expresser line of cmyc tagged proteins?

Any ideas?

Thanks for the help.

Oswaldo



________________________________________________________________

Oswaldo da Costa e Silva, Ph.D.

Institut f=FCr Allgemeine Botanik
Universtit=E4t Hamburg
Ohnhorststr. 18
22609 Hamburg
=46.R. Germany

Tel.:  + 49-40-822 82 391
=46ax.:  + 49-40-822 82 503
Email: ocs at botanik.botanik.uni-hamburg.de








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