In this case, pGEX4T3 vector also expressed inserts constitutively.
<http://www.biovisualtech.com/bvplasmid/pGEX-4T-3.htm>http://www.biovisualtech.com/bvplasmid/pGEX-4T-3.htm
Not quite. pGEX vectors carry a copy of lacI^q, resulting in very low leak
expression from PTAC promoter. Definitely lower than the relatively leaky
T7 promoter in pET20, which does not express laqI^q. Switching to a plasmid
from higher pET series that do have laqI^q (e.g. pET31) should reduce the
leakiness.
I also thought that target protein may have tocicity on host strain.
But, why pGEX4T3-target protein was not show the same phenomena.
Now, we are trying to change the host BL21(DE3) to Rosetta(DE3) and
BL21(DE3)pLysS.
pLysS should help with repression under non-inducing conditions.
If it is true toxicity issue, you might need to switch to plates with
synthetic medium that does not contain any traces of lactose.
- Dima
