Why, at 3, 4.5 , 6, 18 hours after IPTG induction, do we harvest the target protein from E.coli? Is there any reason for this treatment?
Using different times for harvesting the proteins samples from E. coli is a part of standardization process for protein extraction. The reason is that we need maximum amount of protein production in the minimum amount of time. The protein synthesis in E. coli gets saturated after a particular time. To find out that time, this exercise is done. once we know the time of saturation, we just limit our experiment to that time. Say, you are getting same amount of protein after 4.5 hours of induction and there is no increase after that. Then while doing the experiment you can just harvest the protein at 4.5 hours instead of waiting till 18 hours or so. Because after that the concentration of protein is going to be same. Sometimes, solubility of protein also matters. So, taking a time point where protein is soluble and in maximum amount is the best choice and to find out that time, this is done.
The optimum concentration of inducer and the duration of induction is protein specific. You may start with a middle range; 1mM IPTG and 3 hr.. But still you can play around this to find out a point where you get the maximum yield of your protein with minimum cell lysis....
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