I have a rather basic question regarding protein expression (T7 based) in E. coli (I use strains Rosetta and BL21AI, if that matters). I usually induce at OD600 = 0.6, lower the temperature to 18 °C and express overnight (if I express at higher temperature, I have some issues with protein solubility).
OK, so this approach works reasonably well, but I\’m looking for possible improvements regarding some tricky candidates. Recently I thought about switching to TB medium (from LB) for those cases. TB allows me to reach higher ODs thus increasing the potential protein output for a given culture volume under non-time-limited incubation (overnight = culture reaches saturation), if I understand everything correctly.
Now, as I understand the increased buffer capacity and additional carbon source (glycerol) aids in reaching high ODs, and I assume cells grow to higher ODs because reaching the stationary growth phase is delayed. So this means the log phase is longer, right? If that is correct, I assume that I could as well induce at higher OD600 than the standard 0.6, as ”longer log phase” would mean that the ”early log phase” which is the sweet-spot for induction would be extended as well.
Is my reasoning correct and has anybody any practical experience that would be applicable to my case?
Thanks a lot!