T7 promoter strength - (Sep/20/2007 )

Dear all,

will reduction of T7 promoter strength overcome protein expression problems due to codon usage?
If so, how can I reduce the transcription level of my proteins? Take less IPTG (f.ex. 0.1 mM instead of 1 mM)?

Thanks,

chalet2

Hi Chalet,

Reducing the promoter strength might help with codon usage issues, and lowering the IPTG concentration as you suggested is the way to do it. You could also try growing the cells at a lower temperature, which will slow things down and maybe help the cells to cope or you can try expressing in one of the Rosetta strains.

There are also some cells (like BL21 pLys or something) that have a T7 inhibitor.
Sometimes they work better if you have problems with the amount of expressed protein.