S.O.S. PROTEIN OVEREXPRESSION!!!!!

[Jorge.Hernandez at bota.unine.ch]

Hello netters!

I'm trying to overexpress two plant proteins in E.coli. I use the
expression vector pET3d and BL21(DE3) as E.coli strain. Both cDNAS
are cloned on the same strand having each a ribosome binding site (dicistron). 
Thus, they are expected to be co-transcribed but translated separately 
cf. the following scheme:


T7 Promoter !   RBS               RBS
             !  /  protein  1     /      protein 2
            5'===--------------===---------------- 3'
                          
                         -------->
                        TRANSLATION

The problem:
Protein 2 is expressed but not protein 1. I use polyclonal antibodies
to monitor expression of each gene.

The question:
Can anyone help me to explain why protein 1 is not expressed?
I would appreciate all suggestions about.
Thanks in advance,

Georges,

E-mail: Jorge.hernandez at bota.unine.ch
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