IPTG: BL21 vs BL21(DE3)

Dominic-Luc Webb molmed domweb at mbox.ki.se
Wed Oct 17 12:23:04 EST 2001


It was proposed by someone on this list that IPTG
should halt growth of BL21(DE3) lysogens transformed
with a T7 plasmid (I am using pCMX with a non-toxic
target gene). Given what I have read in the
literature (i.e., Studier), this makes good sense. I
have not been getting very good protein expression and
did some investigating into the cause. One parameter
I examined was inability to grow in the presence of
IPTG.

I transformed the pCMX into several strains and
did growth experiments with 3 hrs, 37C +/- 1 mM IPTG
induction. Only BL21(DE3 and BL21(DE3)pLysS should
have chromosomal copies of the T7 RNApol. To my
surprise, I found that BL21 without T7 RNApol seems to
be distinct from several other E coli strains in having
growth that is sensitive to IPTG. The BL21 strains are
control strains from Novagen. Assuming, the E coli are
in fact as labelled by Novagen, it appears that lack of
growth after IPTG induction cannot be taken as an indication
that there is a functional DE3 lysogen in this strain.
I got the following A600 absorbance readings and would
deeply appreciate any comments (absorbance was roughly
linearly related to relative number of cells):


Time 0 (start)
BL21             .014
BL21(DE3)        .011
BL21(DE3)pLysS   .020
DF(6)            .011
DF(4)            .010
XL1_blue         .013


3 hrs w/o IPTG
BL21             .292
BL21(DE3)        .278
BL21(DE3)pLysS   .212
DF(6)            .107
DF(4)            .096
XL1_blue         .121


3 hrs 1 mM IPTG
BL21             .043
BL21(DE3)        .037
BL21(DE3)pLysS   .123
DF(6)            .105
DF(4)            .099
XL1_blue         .137


Note: DF strains are metabolically compromised and
are expected to grow slowly.



Cheers,


Dominic-Luc Webb, doktorand


Lab:
Department of Molecular Medicine
Endocrinology and Diabetes Unit
Rolf Luft Center for Diabetes Research
Karolinska Hospital L3
S-17176 Stockholm
Sweden
Tel: Int+46-8-517-74829
Fax: Int+46-8-517-79450
Internet Email: dominic at enk.ks.se




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