Selective Breeding of a Genome-Scale Metabolic Network

Emory University, Atlanta GA

Investigators

Abstract

Intellectual Merit: The metabolic pathways of any particular cell form a single interconnected network. The Investigator's goal is to understand how this network evolves in response to the changing needs of the organism. He and his team have developed a model system that streamlines laboratory investigations of adaptive genome evolution. They expressed the lux operon of Photorhabdus luminescens, which encodes an anabolic pathway producing visible light, in Acinetobacter baylyi ADP1, which naturally exchanges and recombines its chromosomal DNA as it grows. The model system has an active natural recombination system that enables an efficient microbial breeding strategy and at the same time, permits the selection of gene-based metabolic traits that contribute to the production of visible light. The Investigator proposes to direct the evolution of strains that produce even more light, and to investigate the most hypermorphic strain by genome sequencing, microarray analysis and biochemical studies of the evolved proteins. This work will show how evolutionary tradeoffs can optimize the flux through a model anabolic pathway. These lessons could be applied to increase the biosynthetic yields of biofuels, materials, and other economically valuable compounds. Broader Impact: Many prospective molecular biologists lack the financial resources to conduct research. The cost of the basic equipment necessary to transform Escherichia coli (centrifuge, 80° C freezer) imposes a formidable entry barrier. A. baylyi is naturally competent, so it can be transformed without expensive hardware. The Investigator's team has created a novel expression vector for this organism, and has written software that shows users how to combine DNA fragments in overlap PCR reactions. A determined high school student or hobbyist could use these tools to create and express novel genes in minimally equipped home laboratories. This study will inspire novice scientists to design and conduct their own experiments, and eventually to seek formal training.