Hi everyone. I am trying to express and purify a recombinant antimicrobial peptide using as a host cell E. coli BL21 (AI) (employing L-arabinose and IPTG). Because this peptide can be toxic to cells, I want to know if induction is appropriate when DO600 reaches half of the logarithmic phase (0.4) or if it is better to do so when greater biomass is achieved and stabilized (stationary phase ). Thanks in advance.
Hi there,
As the expected effect is toxicity on cells I would suggest to go for a strong induction at high cell density in order to optimize the yield of production on a very short laps of time... But the fact is that heterologous expression is strongly empirical therefore only experiments will tell you what are the best conditions for your specific case.
Hi there,
As the expected effect is toxicity on cells I would suggest to go for a strong induction at high cell density in order to optimize the yield of production on a very short laps of time... But the fact is that heterologous expression is strongly empirical therefore only experiments will tell you what are the best conditions for your specific case.
As Dominique suggests, your best bet is to use culture conditions that afford the highest cell density, so that you can get the highest biomass while still in the growth phase. This means a rich medium such as 2YT and a good aeration. With these conditions, a non-induced culture will reach OD >5, so you could easily wait until OD 1-1.5 before induction
Is the protein cytoplasmic or periplasmic, then a pilot studies will help you decide the best density for induction, each protein expresses in different ways.. Pierre and Dominique's suggestions are also of great importance..
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