S125: Developing a genetically stable and tractable strain for scaling up 1,4 butanediol production in E. coli

Biochemical processes fundamentally differ from chemical processes because of the self-replicating nature of the catalyst: the living aspect, the microbe.  Each genetic manipulation requires the microbe to self-replicate, perhaps 30 times.  Coincidently, scaling up from 1 milliliter to 1 M liters requires a similar number of cell divisions; a culture vial photographed next to a commercial bioreactor would look much like an E. coli cell photographed next to a culture vial.  So, in the sixty or so generations between picking a colony and harvesting a commercial bioreactor, some change—unintended and untracked—is practically guaranteed.  Rational approaches facilitate coping with this problem.  Recently, Genomatica engineered E. coli to produce 1,4 BDO and to successfully tolerate the generational distance between the design lab and the commercial fermentor.  The training wheels, namely IPTG-inducible systems and antibiotics, were necessarily stripped away as designs matured.  Stability stress tests highlighted unstable sequences and provided clues to selection pressures introduced into the system by constitutively expressed genes.  Despite the occurrence of inconsequential fluctuations in the genome, scale-down experiments under fermentation conditions showed that the catalyst would likely complete the task in much the same form in which it began.