Best basic parts – psbAI promoters (tertiary headers)
For this Project, the promoters were considered as the best or the most important basic parts. For the expression of RubisCO’s genetic circuits the constitutive psbA promoters were chosen because of their light responsive properties and their strength of expression in Synechococcus sp.
These promoter’s expression can be induced rapidly at different wavelengths and intensities, which facilitates the control of the gene expression without the need for the addition of any inducer to the culture media, instead, the regulation is carried out by a physical stimulus. Following this logic, we designed two modules for a two-stage culture strategy: in the first one, biomass carbon uptake is increased and once the nitrogen source is depleted from the culture media and stress responses from the cyanobacteria are activated, free fatty acids biosynthesis is induced.
Phase 1: psbAI promoter (Tertiary header)
The transcripts from psbAI promoter originate under low light intensity conditions (125 mol photons m-2 s-1). The carbon fixation and biomass production phase are under the control of this promoter (BBa_K754000).
The expossure of the cells at high light conditions (750 µmol photons m-2 s-1) decreases the psbAI promoter strength while the expression of the genes regulated by the promoters psbAII and psbAIII increases. Also, it has been shown that low blue light conditions activate the psbAII and psbAIII promoters [1].
For this reason, we designed a hybrid psbAII/III promoter that allows a high transcription rate under high blue light intensity to increase the FFAs production after an increased carbon uptake by the cells.
These designs allow to regulate the expression of our gene circuits with an easy and effective mechanism.
[1] Mulo, P., Sicora, C., & Mari, E. (2009). Cyanobacterial psbA gene family: optimization of oxygenic photosynthesis. Cellular and Molecular Life Sciences, 66, 3697-3
