Hello everyone, I have a question about recombinant e.coli culture.
At my last experiments, I found that my recombinant e.coli expressed target protein very well.
so I wanted to increase their productivity, I changed medium composition and added carbon source. Here is medium composition,
carbon source 10g, yeast extract 5g, KH2PO4 13.5g, MgSO4 1.4g /L
I used glucose as a carbon source and I cheaked remaining glucose for an hour, but my r-E.coli did not use glucose a lot.
( I cultured it for 6hrs. first glucose percentage was 1% and it was 0.83% after 6hrs)
Normally they prefer glucose, so I know that they use most of glucose in 4hr. but why my e.coli does not like glucose? For reference, my protein is outer membrane protein and both protein and e.coli have toxicity.
Does anyone have simmilar experience before?
Does ur e coli strain have all the components of PTS? R U sure ur insert does not affect EII b-c of the PTS?
May be unrelated but another way to increase yields that often (but not always) works is to switch to some of those optimized commercially available auto-induction medias. Is that what you're trying to create yourself? In my experience it just doesn't seem to work for some proteins.
I am wondering which pH your medium has. The KH2PO4 will cause an acidic pH, requiring adjusting with NaOH.
Generally, when the medium composition is changed, the effect on the product yield is hard to predict. Design of Experiments (DoE) schemes are often successful for media optimization.
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