High throughput screen for cells with high extracellular metabolite consumption--secretion rates using microfluidic droplets

Metabolic engineering has contributed significantly to the improvement of strains for the industrial production of various compounds. Traditionally, enzymatic steps closely associated with the product-forming pathway have been engineered to prune side reactions and eliminate kinetic bottlenecks. Other, so-called distal genes may also impact production in a profound way due to (often unknown) kinetic and regulatory effects. Inverse Metabolic Engineering (IME) emerged as an approach to identify such distal genetic factors. IME employs combinatorial methods whereby libraries are constructed harboring random genomic variants of the host or other strains, cells with superior properties are selected, and genetic inserts impacting the superior phenotype are characterized. While many strategies can be deployed in library construction, broad applicability of IME to strain improvement for overproduction of secreted metabolites is severely limited by the availability of high-throughput methods for selecting strains with significantly improved metabolite secretion or uptake rates. As soon as a metabolite is secreted, its association with the cell that secreted it is lost. Thus, each clone must grow in a separate environment to allow for the measurement of clone-specific metabolite concentrations. Furthermore, since many mutant libraries are large (=/>104 unique clones), a high throughput screening platform must be used to culture and measure each unique clone.

Department dc:contributor.department

Massachusetts Institute of Technology. Dept. of Chemical Engineering.

Grantor dc:publisher

Massachusetts Institute of Technology

Year dc:date.issued

2009

Author dc:creator

• Wang, Benjamin L. (Benjamin Lu chen)

Advisor dc:contributor.advisor

• Gregory N. Stephanopoulos.

• Chemical Engineering.

Statement dc:rights

• M.I.T. theses are protected by copyright. They may be viewed from this source for any purpose, but reproduction or distribution in any format is prohibited without written permission. See provided URL for inquiries about permission.

Licence dc:rights.uri

• http://dspace.mit.edu/handle/1721.1/7582

Language dc:language.iso

eng

Handle dc:identifier.uri

http://hdl.handle.net/1721.1/51674

OAI identifier oai:identifier

oai:dspace.mit.edu:1721.1/51674