Poly(3-hydroxybutyrate-co-4-hydroxybutyrate) Production By Halomonas TD01 in Continues and Open Fermentation Process with Low Sodium Concentration

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Poly(3-hydroxybutyrate-co-4-hydroxybutyrate) [P(3HB-co-4B)], which is synthesized by a variety of bacterial strains, shows different mechanical properties with various 4HB ratios. So far, the industrial production of P(3HB-co-4HB) is generated from structurally related carbon sources, such as 1,4-butanediol or γ-butyrolactone. And the high cost of the fermentation process and raw materials are the disadvantages for the production and application of P(3HB-co-4HB). Now, a pathway for P(3HB-co-4HB) production from glucose was successfully constructed in Halomonas TD01, and further genes manipulation will be done to enhance the metabolic flux for higher 4HB ratio (10%-14%). On the other hand, Halomonas TD01 is a strong candidate for PHA production in continuous and open fermentation using seawater instead of fresh water for its high pH toleration. In our case, we can generate 60% PHA content of cell dry weight in shake flasks in MM medium with 30 g/L glucose only, and the 4HB ratio of P(3HB-co-4HB) copolymer production is around 1.8%. And the highest cell dry weight was more than 61 g/L and the 4HB ratio was about 1.3% in 1 litter continuous and open fermentation trial. At the same time, we are making effort to knock-out gabD gene to enhance the 4HB ratio, and three copies were successfully knocked out. Furthermore, we have brought down the NaCl concentration from 60 g/L to 7.5 g/L in MM medium for Halomonas TD01 to grow with some genes deletion, which can dramatically reduce the cost and corrosion of the bioreactor. All the above show the great effort and potential for the industrial low cost P(3HB-co-4B) production in the future.