Expression of recombinant protein in E. coli can be an efficient and useful technique to produce large quantities of protein quickly. The same plasmid that can be used in small-scale productions to produce microgram quantities of protein can also be scaled up to produce milligram and even gram quantities of protein. However, individual expression constructs may have very specific growth requirements for optimal protein expression.
One requirement that is often temperamental is the point in the bacterial growth cycle at which induction of protein expression occurs. Most constructs express optimally if the induction occurs during mid-log phase when the cells are dividing exponentially. The difficulty here lies with the observation that different expression constructs result in the bacteria potentially possessing quite different growth profiles. In order to assure that the cells are indeed undergoing logarithmic growth, most protocols suggest inducing when the OD600 = 0.6 or above. Unfortunately, because of the varied growth characteristics of cultures expressing different constructs, determining when the cells are at an OD600 of 0.6 can often become a time consuming and labor intensive task. In addition, some labs may not have the necessary equipment to directly measure OD600 rendering this task even more difficult.
Novagen offers a unique solution to this problem, media that allows the cells to grow to a high density and then automatically induces the cells when they have reached the correct growth state without the need to add IPTG. The theory behind the product is that by using a fully defined complex media, the cells are able to grow to saturation while repressing expression from the expression construct and then spontaneously induce high level expression of the target protein. This all takes place without the need to monitor cell growth or add inducing agents like IPTG.
In theory, this is a marvelous system which reduces the labor involved in expressing recombinant protein in E. coli. In practice, unfortunately, the results, while satisfactory, do not live up to the advertising. We have used this system to express protein from several IPTG inducible plasmids, coding for recombinant proteins ranging in size from ~25 kDa to 45 kDa, but have not used this system for expressing larger proteins. While the convenience of simply inoculating the culture and letting it go is wonderful, in our hands, yields were consistently lower than when the same construct was grown to OD600=0.6 and induced with IPTG. The yields were not dramatically different however, we saw yields reduced by as much as two-fold. This difference was enough to require a second induction in order to produce sufficient protein for our experiments.
With this caveat taken into consideration, this system offers a very attractive option for labs that do not routinely express proteins. The formulation is simple, the protocol easy and the yields adequate. The price also makes this system quite attractive as it most likely can be purchased for approximately the same cost as typical media components and induction reagents.