A Logic NAND Gate for Controlling Gene Expression in a Circadian Rhythm in Cyanobacteria.

To enable circadian control of gene expression in cyanobacteria, we constructed a genetic logic gate (NAND) using orthogonal promoters via modular CRISPR interference. The NAND gates were tested in Synechococcus elongatus PCC 7942 using a fluorescent reporter. The NAND gate dynamics were characterized based on the affinity of the dCas9 complex to the output element. Upon connecting tight gene repressions with the circadian promoter (the purF gene; peak expression at dawn), inversed peak expressions were obtained as an output of the NAND gate although the retroactivities were shown in the ON and OFF states. A dark-responsive genetic element of the NAND gate was also expanded to an AND gate in S. elongatus PCC 7942. These cyanobacterial NAND and AND gates could facilitate the control of gene expressions in dynamic metabolic engineering technologies, thereby enabling the cyanobacteria to serve as biosolar cell factories.