A promoter–RBS library for fine-tuning gene expression in <i>Methanosarcina acetivorans</i>

Methanogenic archaea are potent producers of the greenhouse gas methane and thus contribute substantially to global warming. Under controlled conditions, these microbes can catalyze the production of biogas, which is a renewable fuel, and might help counter global warming and its effects. Engineering the primary metabolism of <i>Methanosarcina acetivorans</i> to render it better and more useful requires controllable gene expression, yet only a few well-characterized promoters and RBSs are presently available. Our study rectifies this situation by providing a library of 33 different promoter-RBS combinations with a 140-fold dynamic range in expression strength. Future metabolic engineering projects can take advantage of this library by using these promoter-RBS combinations as an efficient and tunable gene expression system for <i>M. acetivorans</i>. Furthermore, the methodologies we developed in this study could also be utilized to construct promoter libraries for other types of methanogens.