Hello everybody, Sorry for an offtopic question. I am trying to express a protein in M9 minimal media for Selenomet incorporation. When grown in LB this protein expressed very well and got good crystals. Diffraction was upto 2 A. I am having a hard time expressing the same protein in Minimal media. It took nearly 24 hours for the OD600 to reach ~ 0.4 before inducing with IPTG in minimal media and eventually got no protein expression. It looks like the cells are not growing or taking very long to grow. The cell line I am using is RossettaBlue (DE3) and the plasmid is pET19b based (Novagen).
It expressed poorly in BL21 (DE3) when grown in LB and thus decided to use RosettaBlue (DE3). It worked very well in LB, but having a hard time while expresing the same in minimal media using Rosetta Blue. Has anybody tried expression in minimal media using Rosetta Blue cell line? I am planning to try overnight induction. Any suggestions would be greatly appreciated. Thanks, Manish ************************************************* Manish B. Shah, PhD. Postdoctoral Fellow Hauptman-Woodward Medical Research Institute 700 Ellicott Street Buffalo, NY 14203. *************************************************
