A promoter library for tuning gene expression in Cupriavidus necator under autotrophic conditions
This study aimed to develop a promoter library for Cupriavidus necator that functions under autotrophic conditions. C. necator was cultured under both heterotrophic and autotrophic conditions, and comparative transcriptome analysis was performed to identify genes and operons specifically upregulated under autotrophic conditions and those constitutively expressed. The upstream sequences of the candidate genes and operons were examined to identify their promoter regions. We established a promoter evaluation system based on colorimetric measurement of beta-galactosidase enzyme activity in C. necator. Utilizing this system, we successfully identified eight and three promoters that specifically upregulate the downstream beta-galactosidase under autotrophic conditions and three promoters that constitutively express the downstream beta-galactosidase under both autotrophic and heterotrophic conditions. This study represents the first development of a promoter library that functions in C. necator under autotrophic conditions, which will contribute to the advancement of efficient biomanufacturing processes using CO2 as the feedstock.