Abstract:Development of “liquid sunshine” could be a key technology to deal with the issue of fossil fuel depletion. β-caryophyllene is a terpene compound with high energy density and has attracted attention for its potential application as a jet fuel. The high temperature and high light-tolerant photosynthetic cyanobacterium Synechococcus elongatus UTEX 2973 (hereafter Synechococcus 2973), whose doubling time is as short as 1.5 h, has great potential for synthesizing β-caryophyllene using sunlight and CO2. In this study, a production of ~121.22 μg/L β-caryophyllene was achieved at 96 h via a combined strategy of pathway construction, key enzyme optimization and precursor supply enhancement. In addition, a final production of ~212.37 μg/L at 96 h was realized in a high-density cultivation. To our knowledge, this is the highest production reported for β-caryophyllene using cyanobacterial chassis and our study provide important basis for high-density fuel synthesis in cyanobacteria.