Genetics of Unstudied Thermophiles for Industry
Thermophilic organisms hold great potential for industry due to their numerous advantages in biotechnological applications such as higher reaction rate, higher substrate loading, decreased susceptibility to reaction contamination, energy savings in industrial fermentations, and ability to express thermostable proteins that can be utilized in many important industrial processes. Bioprospecting for thermophiles will continue to reveal new enzymatic and metabolic paradigms with industrial applicability. In order to translate these paradigms to production scale, routine methods for microbial genetic engineering are needed, yet remain to be developed in many newly isolated thermophiles. Major challenges and recent developments in the establishment of reliable genetic systems in thermophiles are discussed. Here, we use a hyperthermophilic, cellulolytic bacterium, Caldicellulosiruptor bescii, as a case study to demonstrate the development of a genetic system for an industrially useful thermophile, describing in detail methods for transformation, genetic tool utilization, and chromosomal modification using targeted gene deletion and insertion techniques.
© Springer Science+Business Media, LLC, part of Springer Nature 2020. First Online: 28 July 2020. We thank Gina L. Lipscomb for sharing C. bescii transformation protocols. Funding was provided by the BioEnergy Science Center (BESC) and the Center for Bioenergy Innovation (CBI), from the U.S. Department of Energy Bioenergy Research Centers supported by the Office of Biological and Environmental Research in the DOE Office of Science. This work was authored in part by Alliance for Sustainable Energy, LLC, the Manager and Operator of the National Renewable Energy Laboratory for the U.S. Department of Energy (DOE) under Contract No. DE-AC36-08GO28308. The views expressed in the article do not necessarily represent the views of the DOE or the U.S. Government. The U.S. Government retains and the publisher, by accepting the article for publication, acknowledges that the U.S. Government retains a nonexclusive, paid-up, irrevocable, worldwide license to publish or reproduce the published form of this work, or allow others to do so, for U.S. Government purposes.