The Construction Of Pinene Synthesis Pathway In Synechococcus Sp.PCC 7002

The excessive use of fossil fuels not only causes energy shortage,but also causes a variety of environmental problems,and the use of bio-fuels is one of the effective ways to ease the energy crisis and environmental degradation.At present,the use of microbial synthesis platform to convert those sufficient and cheap raw materials into a variety of bio-fuels is a hot spot of bio-fuel research.However,it is relatively scarce for the development of high-energy-density biofuel.Pinene,an important plant secondary metabolites,is mainly found in plant essential oil,whose dimer has a similar higher energy density with JP-10.Nowdays,the heterologous synthesis of pinene has been successfully achieved in E.coli and Corynebacterium glutamicum.Cyanobacteria,by contrast,has become an important photosynthetic autotrophic microbial platform for heterologous synthesis of plant secondary metabolites with its outstanding photosynthetic capacity,rapid growth rate,ease of genetic manipulation as well as high cell density.In this paper,we constructed an integrative plasmid containing the pinene synthase gene fragment,and the pinene synthase gene was integrated into the cyanobacterium genome by double homologous recombination using the homologous fragment of pAQ1,an endogenous plasmid of Synechococcus sp.PCC 7002.Resistant strains were identified by PCR and protein immunoblot analysis,and the target strains were identified at 30?,250?Em^-2s^-1 and 1%CO₂.The highest yield of pinene was 41?g/L.Under the condition of changing CO₂ concentration,temperature and light intensity,the highest yield was obtained which has reached 169?g/L.In this paper,the synthesis of pinene in Synechococcus sp.PCC 7002 for the first time has greatly expanded the application prospect of Synechococcus sp.PCC 7002.