We are aware of an issue with certificate availability and are working diligently with the vendor to resolve. The vendor has indicated that, while users are unable to directly access their certificates, results are still being stored. Certificates will be available once the issue is resolved. Thank you for your patience.

(711g) Rapid Evolution of Regulatory Elements for Multiplexed Combinatorial Metabolic Engineering

Xu, P., MIT
Stephanopoulos, G. N., Massachusetts Institute of Technology

Optimizing microbial cell factory to produce targeted chemicals often requires the expression of multiple enzymatic steps. It is desirable to coordinate the expression of these genes to prevent the accumulations of toxic intermediates/byproducts and maximize the carbon flux towards the target pathway. To meet this challenge, recent advances in metabolic engineering have spurred great interests to explore precise and combinatorial approaches to control and optimize biosynthetic pathways for production of fuel and pharmaceutical molecules. Resembling of the engineering principle, most of the current combinatorial pathway optimization follows a rather intuitive framework – an iterative cycle of design, characterization, construction and testing. Despite great success to improve the production of an array of metabolites, the current approach suffers from tedious trial-and-error work that obviously restricts our ability to realize the full potential of a specific biosynthetic pathway. To bridge this gap, we have invented an evolutionary approach that efficiently incorporates single strand regulatory DNAs (ssDNAs) and rapidly tuned the expression of multiple genes at the same time. As a case study, we have simultaneously evolved the promoter, operator and terminator regions of a five gene pathway and resulted in significant titer improvement. Combining with sophisticated DOE (design of experiments) principles, this approach opens up new avenues for quantitative multiplexed metabolic engineering.