Metabolic Engineering and the Synthetic Biology Toolbox for Clostridium

The Clostridium genus contains a diverse range of Gram-positive, sporulating, obligate anaerobes that have been of historical biotechnological interest due to their acetone–butanol–ethanol (ABE) solvent production. Within the last few decades, interest has grown in the Clostridium spp. capable of consuming a wider variety of feedstocks, which include gaseous and renewable biomass sources. Additionally, attenuated pathogens have been of interest as potential therapeutics. The fruition of the genus's great promise has been limited by the slow progress in genetic engineering and synthetic biology methods and tools, relative to workhorse organisms such as Escherichia coli . Recent advances in these areas, not least of which include CRISPR-based tools, renew the promise of metabolic engineering for a broad range of feedstock consumption and production of chemicals. In this chapter, we describe the current state of engineering in the Clostridium genus by describing efforts and continued challenges in directed evolution, the use of systems biology for greater understanding, methods for performing genomic editing, and the expanding library of genetic parts. These new capabilities and tools have expanded the number of species that are able to be engineered for biotechnological purposes, increased the throughput of genetic studies, and expanded the range of products made from Clostridium.