my problem in protein expression in pre-culture step that I already prepared it in 20 ml LB and added 20 micro gram/ml Chloramphenicol,50 micro gram/ml ampicillin,5ng/ml tetracycline and 0.5 mg/ml L-arabinose. using cell stock incubate at 37 c for 5 hours but no growing I wonder the problem because of incubation time or cell stock has a problem ??
How exactly are you inoculating that 20 mL starter culture? I wouldn't expect a 20 mL culture inoculated with a single colony (or with a relatively small amount of cells) to exhibit noticeable turbity after only 5 h at 37oC, and certainly not with the metabolic burden of chaperone co-expression and maintaining an additional plasmid. Wait for longer until the culture reaches exponential growth (0.4-0.6 DO would be fine) and then transfer to 15oC and add IPTG to induce expression from pCold.
Hope it helps!
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