Protein expression in E.coli DH5alpha? - (Dec/11/2007 )
I have been trying to use DH5alpha to screen for protein expression, as I can then use the same culture for both the DNA prep and the expression. However, I have noticed that I have abundant expression of my protein under non-induced conditions. I am using the pGEX 6P-1 GST fusion vector, from GE Healthcare - which carries the lacIq repressor, and which should, in theory, not allow any protein synthesis in the absence of IPTG. I am wondering if anyone have any comments on why I observe protein expression without addition of IPTG?
William Studier (of pET expression fame) showed that many rich media based on casein digests actually have sufficient trace lactose to induce expression. His paper in 2005 (Prot Exp Purif 41:207-234) discusses many of these issues in detail. Being the good academic that he is Studier has handed out his recipes for media to many researchers; so many, in fact that hee is about to prepare a final, hopefully definitive, set of conditions. While these may apply especially to the pET (that is the T7 RNA pol system) vector, there will be plenty of interest for other expression systems.