Hello. I want to isotopically label the my protein, and I express it in BL21 RIPL using M9 minimal media. It grows extremely slow.
I inoculate the colony from a fresh M9 plate into 5ml M9 medium as the preculture. But the cell culture is still clear after overnight incubation at 37 degree.
It seems some nutrient is missing. Is it vitamin B1 (thiamine) missing? Could anyone tell me the nutrient requirements based on the genotype (please see below)?
E. coli B F– ompT hsdS(rB – mB – ) dcm+ Tetr gal λ(DE3) endA Hte [argU proL Camr ] [argU ileY leuW Step/Specr ]
The recipe I am using is:
5*M9 salt: KH2PO4, 15 g/L NaCl, 2.5 g/L Na2HPO4, 33.9 g/L NH4Cl, 5 g/L
For 1 L M9 medium:
5*M9 salt: 200ml
1M MgSO4: 2ml
1M CaCl2: 0.1ml
0.5g/L FeSO4. 7H2O: 1ml
20% Glucose: 20ml
Thanks in advance.
Best wishes
Jingjing
Also check https://www.chem-agilent.com/pdf/strata/230240.pdf
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