Design and Reconstruction of Regulatory Parts for Fast-Growing <i>Vibrio natriegens</i> Synthetic Biology

Abstract

The fast-growing <i>Vibrio natriegens</i> is an attractive robust chassis for diverse synthetic biology applications. However, <i>V. natriegens</i> lacks the suitable constitutive regulatory parts for precisely tuning the gene expression and, thus, recapitulating physiologically relevant changes in gene expression levels. In this study, we designed, constructed, and screened the synthetic regulatory parts by varying the promoter region and ribosome binding site element for <i>V. natriegens</i> with different transcriptional or translational strengths, respectively. The fluorescence intensities of the cells with different synthetic regulatory parts could distribute evenly over a wide range of 5 orders of magnitude. The selected synthetic regulatory parts had good stability in both nutrient-rich and minimal media. The precise combinatorial modulation of <i>galP</i> (GalP = galactose permease) and <i>glk</i> (Glk = glucokinase) from <i>Escherichia coli</i> by using three synthetic regulatory parts with different strengths was confirmed in a phosphoenolpyruvate:carbohydrate phosphotransferase system with inactive <i>V. natriegens</i> strain to alter the glucose transport. This work provides the simple, efficient, and standardized constitutive regulatory parts for <i>V. natriegens</i> synthetic biology.