I want to know in pET28a/BL21DE3 and also in pCold TF/BL21DE3 for how much time (hrs) after induction with IPTG culture should be kept and at which

 

temperature so that soluble aggregates should not form? I kept at 18 degree for 18 hrs after induction in pET28a and at 15 degree for 24 hrs but there is formation

 

of soluble aggregates in both as detected by Native PAGE . I read that if kept for long time protein forms aggregates or come in inclusion bodies.

 

 

 

In my limited experience, each situation is different. Therefore, you will probably need to experimentally determine conditions. If your protein has a prosthetic group, sometimes the growth medium needs to be enriched a chemically related species. Some proteins, however, refold nicely from inclusion bodies.

ok sir !!!! thanks!!!!!!!!