two-hybrid, detection fusionprotein?

reinhilde at lmb1.rug.ac.be reinhilde at lmb1.rug.ac.be
Thu May 18 02:16:58 EST 1995


From: reinhilde at lmb1.rug.ac.be
Subject: two-hybrid, detection fusionproteins?
Date: Wed, 17 May 1995 16:49:26 GMT

Hello,

I made a fusionprotein with the GAL4 DNA-binding domain in the pAS2 vector 
and on Westernblot I could detect the product on the expected 
migration-distance. The in-fase-fusion was indeed confirmed by sequence 
analysis. The immunodetection was achieved by using an anti-GAL4-DNA 
BD-antibody. Unfortunately I have never been able to detect my protein of 
interest (C-terminal of the fusion) although I tryed several specific 
antibodies that should recognize it. Is there something wrong with the 
protein-folding in the yeast cells?

Does anybody succeded in detecting the second part of the BD-fusion by 
specific immuno- or other detections? 
Please let me know if you did, how big the protein is and if it is (in natural 
conditions) an intracellular protein or not.

Thanks for your time and answer...

Reinhilde Schoonjans,           tel. : (32)9/264.53.17
Lab Molecular Biology           e-mail: Reinhilde at lmb1.rug.ac.be
University of Ghent
Belgium

  




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