expression with BL21(DE3)

Dr. Peter Gegenheimer PGegen at UKans.edu
Fri Nov 14 18:58:16 EST 1997


On Wed, 5 Nov 1997 19:19:29, moffatt at SCIBORG.UWATERLOO.CA (Barbara 
Moffatt) wrote:

> Hello:
> 
> I have a colleague that wishes to transform a plasmid library created from
> that Stratagene lambda zap (unizap) vector into BL21(DE3) for expression
> from the T7 promoter. Can anyone comment on the expression levels from a
> non pET vector, using a BL21(DE3) host? Novagen says they don't guarantee
> it and I only have experience using pET vectors with this host.
> Thanks in advance for your assistance.
> 
> Barb Moffatt
> moffatt at sciborg.uwaterloo.ca

I'm surprised anyone has every gotten results with this system. The 
BL21(DE3) cells provide *regulated* expression of 
T7-promoter-controlled genes **ONLY** from a low-copy-number plasmid
like pBR322. High-copy plasmids based on pUC (such as the 
pBS/Bluescript) vectors provide constitutive expression of the T7 
promoter, *unless* the plasmid also carries a functional lacI gene. 
(The reasons for this should be obvious.) SO, one can expect 
acceptable results using a T7 promoter plasmid with lacI, or if one 
is interested in expression only of genes whose products do not slow
the growth of E. coli. 

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