Dear ccp4bb,
We used to routinely overexpress a number of mammalian proteins to
inclusion bodies in E.coli. Lately however we get from time to time
smeared and small or no pellets after spinning down the cultures (lysed
I assume).
I have read (with some horror) previous threads pointing at phage
contamination and toxic proteins. I am doubtful about the phages as the
problem is not expanding and very frustratingly it appears seemingly
random, coming and going. Finally the one time I checked I did not see
any plaque formation on a e.coli plate tested with a lysed culture.
We have expressed the proteins before many times to inclusion bodies
without any problems. I still could imagine that they could be toxic
when soluble. But why would suddenly happen since we did not change any
conditions that we are aware of?
It would be really great to get some of your ideas on what else could
cause this kind of problem (IPTG, aeration, leftovers from glassware
cleaning, media) or if I missed something with respect to the phages.
Many thanks for your time!
hannes uchtenhagen
--
Hannes Uchtenhagen
Karolinska Institutet
Center for Infectious Medicine (CIM)
Karolinska University Hospital Huddinge, F59
SE-141 86 Stockholm, Sweden
Office: +46-(0)8-524 86981
Mobile: +46-(0)7-36901461
