IPTG induction - (Aug/05/2006 )
i have expressed my gene cloned into a pET23D vector and induced it with two different conc. of IPTG i.e. 0.5mM and 1mM. the protein expression was better i n the 0.5mM IPTG conc. i want to know what is the effect of diffrent conc. of IPTG what forms the basis of the conc. selection as we all know that high conc. of IPTG will lead o cell death?
you use the concentration that gives you the best yield...many proteins are somewhere around 1, so that's where many protocols start...but it's always a ballpark figure
if less gives better expression of your protein...hmm...need a little more detail...I would guess more encourages your protein to form insoluble bodies because too much is made; hence you would lose it during the extraction..although I suppose this is not the only possibility
growing conditions are one of the main limiting factor on a protein production.
So if you medium freshness, culture OD at start were the same, IPTG can be the limiting parameter.
But i don't think it's a limiting factor per se.
Maybe the overinduction leads to a too big quantity of your proteins, and a part of the protein becomes insoluble or unextractible.
Did you check that possibility too ?