Question: Specifically regarding Ampicillin; When growing cells in TB (terrific broth) for protein expression - when should I expect the ampicillin to be gone due to degradation by b-lactamases? (and how much Ampicillin do people add in addition to standard 100 ug/ml)
Background: I have heard rumours that all Ampicillin (100 ug/mL) is degraded within few hours (2-3) due to the enormous amount of b-lactamases secreted by the bacteria.I have also heard that some are not concerned about this, because 2-3 hours is anyway enough for the culture to outcompete anything else that will grow after the ampicillin is gone. On the other hand, some people are also writing that they add more ampicillin every 2-3 hours to maintain selectivity; however, I dont really see the point in maintaining selectivity as long as I have plenty of protein I will anyway lyse the cells and not use them for anything else.
My growth conditions: Grow at 37 degrees till OD600=1 (takes around 3h), then induce with IPTG and grow at 25 degrees for 18h, 200 rpm the entire time.
End comment: I know you can generally change to the more expensive carbenicillin which is degraded slower, but i'm more interested in knowing "how bad" Ampicillin really is for my general knowledge (and I will anyway keep on using it). Also: Any general thoughts or references to literature on this topic, specifically for Ampicillin is appreciated.