A High-Efficacy CRISPR Interference System for Gene Function Discovery in Zymomonas mobilis

Author:

Banta Amy B.12ORCID,Enright Amy L.12ORCID,Siletti Cheta1ORCID,Peters Jason M.1234ORCID

Affiliation:

1. Pharmaceutical Sciences Division, School of Pharmacy, University of Wisconsin—Madison, Madison, Wisconsin, USA

2. Great Lakes Bioenergy Research Center, Wisconsin Energy Institute, University of Wisconsin—Madison, Madison, Wisconsin, USA

3. Department of Bacteriology, University of Wisconsin—Madison, Madison, Wisconsin, USA

4. Department of Medical Microbiology and Immunology, University of Wisconsin—Madison, Madison, Wisconsin, USA

Abstract

Biofuels produced by microbial fermentation of plant feedstocks provide renewable and sustainable energy sources that have the potential to mitigate climate change and improve energy security. Engineered strains of the bacterium Z. mobilis can convert sugars extracted from plant feedstocks into next-generation biofuels like isobutanol; however, conversion by these strains remains inefficient due to key gaps in our knowledge about genes involved in metabolism and stress responses such as alcohol tolerance. Here, we develop CRISPRi as a tool to explore gene function in Z. mobilis . We characterize genes that are essential for growth, required to ferment sugar to ethanol, and involved in resistance to isobutanol. Our Z. mobilis CRISPRi system makes it straightforward to define gene function and can be applied to improve strain engineering and increase biofuel yields.

Funder

U.S. Department of Energy

Publisher

American Society for Microbiology

Subject

Ecology,Applied Microbiology and Biotechnology,Food Science,Biotechnology

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